http://2009.igem.org/wiki/index.php?title=Special:Contributions/Mitch_p&feed=atom&limit=50&target=Mitch_p&year=&month=2009.igem.org - User contributions [en]2024-03-29T15:59:15ZFrom 2009.igem.orgMediaWiki 1.16.5http://2009.igem.org/Team:Alberta/DNAanchorTeam:Alberta/DNAanchor2009-10-22T03:44:12Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#e1e1e1; margin:0 5px;}<br />
.b2f {height:1px; background:#e1e1e1; margin:0 3px;}<br />
.b3f {height:1px; background:#e1e1e1; margin:0 2px;}<br />
.b4f {height:2px; background:#e1e1e1; margin:0 1px;}<br />
.content {background: #e1e1e1;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<br />
<h1>DNA Anchor/Terminator</h1><br />
<br />
<!-- <br />
<h3>Brick Creation</h3><br />
<br />
<p>The use of uracil-containing primers and USER<sup>TM</sup> enzyme mix provides an alternative method for creating 12 base sticky ends that is more versatile than conventional restriction endonucleases. The primers anneal to the A and B regions respectively, as well as ~5bp 3' into the cassette (to increase melting temperature). Bricks cloned into pAB and pBA can be PCR'd up with the universal uracil primers prA1u/prB1u (for pAB), or prA2u/prB2u(for pBA) and treated with USER<sup>TM</sup> mix. The uracil DNA glycosylase (UDG) present in the mix will cleave the uracil base off the DNA backbone. Endonuclease VIII will subsequently cleave the sugar-phosphate backbone at the apyrimidinic site generated by UDG, creating single stranded regions which can be purified away using PCR purification spin columns or gel purification. See <B>Figure 1</B>. </P><br />
<p> A protocol for generating Bytes can be found <a href="https://2009.igem.org/Team:Alberta/Project/ByteAmplification">here</a> and more information on Byte formation can be found <a href="https://2009.igem.org/Team:Alberta/Splasmids">here</a>.<br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/e/e0/UofA09_Bead_Overview_anchor2.png"><br />
<p><B>Figure 1</B>: Showing anchor and terminator fragments and effect of USER<sup>TM</sup> treatment. I SceI site and A ends are highlighted<p><br />
</center><br />
<br><br />
<br />
!--><br />
<br />
<h3>Anchoring System</h3><br />
<br />
<br />
<p>A vital component of the BioBytes method is the use of a biotinylated DNA Anchor in order to allow unidirectional assembly of the Bytes on paramagnetic beads by sequestering the 5' ends of Bytes, leaving only the 3' ends available to bind incomding Bytes. The Anchor itself has three vital components: A 5’ biotinylation, a double stranded DNA (dsDNA) portion that incorporates a release mechanism in order to liberate the construct from the beads, and A or B overhangs to allow Bytes to bind to the Anchor. Our team has considered a number anchor systems, each with their own set of advantages and disadvantages. The three main anchor systems we investigated are summarized below. See <B>Table 1</B>.</P><br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/f/f5/Anchorcomparison.png"><br />
<p><B>Table 1:</B> Overview of three different anchoring systems that were considered for BioBytes. See the section on <B>Anchor Variants and Binding Capacity</B> below for further information on these systems. 5' Biotin (BTN), single stranded DNA (ssDNA), nucleotide (nt), double stranded DNA (dsDNA).<p><br />
</center><br />
<br><br />
<br />
<p>The current BioBytes anchor system utilizes a 5’-biotin which anchors the construct to beads by binding non-covalently, but with great strength, to the covalently linked streptavidin on the surface of the paramagnetic beads. There is also a 5’-15 nucleotide spacer region of ssDNA which facilitates more efficient binding of the Anchor to the bead as the binding pocket of streptavidin is deep and thus a highly flexible ssDNA linker is recommended to allow the biotin to effectively bind into this deep pocket. A 21 bp double stranded portion of the Anchor contains the I-SceI recognition sequence, which when digested with I-SceI produces 4 base overhangs, but also includes four deoxyuracil residues. These uracils are excised by New England Biolab’s USER<sup>TM</sup> system to generate single nucleotide gaps in the top strand. USER<sup>TM</sup> digestion thus effectively destroys the Anchor and produces a 21 base 3’ overhang which becomes important for recircularization of the construct. Finally the Anchor contains the A or B 3’ overhangs complementary to those of the Bytes, allowing their binding to the Anchor. See <B>Figure 1</B>.</P><br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/e/e0/UofA09_Bead_Overview_anchor2.png"><br />
<p><B>Figure 1:</B> pAB and pBA multiple cloning sites with highlighted primers pAB_F/R and pBA_F/R annealing regions.<p><br />
</center><br />
<br><br />
<br />
<br />
<br />
<h3>Termination System</h3><br />
<br />
<p>Once the construct has been completed, i.e. the last Byte has been added, the construct may be released from the beads as is by a simple I-SceI digestion or a USER<sup>TM</sup> digestion, thus yielding a linear construct. If a circular construct, such as a plasmid, is desired then a final "Terminator" piece must be added. This piece is similar in construction to the Anchor, whereby there is a dsDNA I-SceI recognition sequence with four deoxyuracils incorporated into it on one strand, as well as an A or B 3' overhang. The Terminator binds to the last Byte and release is once again achieved by I-SceI digestion or USER<sup>TM</sup> digestion. In either case both the Anchor and Terminator develop sticky ends that are complementary to each other: 4 bases if I-SceI digestion is utilized, or 21 bases if USER is used. USER<sup>TM</sup> digestion is obviously preferred since 21 bp of interaction will form spontaneously and without ligation, and thus transformation of the construct can proceed immediately. See <B>Figure 1</B>.</P><br />
<br />
<h3>Anchor and Terminator Oligo Sequences</h3><br />
<p>The following sequences (<B>Figure 2</B>) are for the oligonucleotides one must order and anneal to generate the full set of Anchors and Terminators. The Anchor_A piece must be used in an Anchor that is meant to bind an AB Byte and Anchor_B for a BA Byte. Terminators containing the Term_A piece will bind BA Bytes, whereas those with a Term_B will bind AB Bytes; remembering that Terminators bind the the 3' end of Bytes and Anchors to the 5' end of Bytes. The Term_Comp and Anchor_Comp sequences are the complementary sequences that anneal to both the Term_A/B and Anchor_A/B pieces respectively to give the 21 bp dsDNA portion of both the Anchor and Terminator. Thus if you want to make an Anchor with an A overhang, you must anneal Anchor_A with Anchor_comp, etc.</p><br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/3/31/Anchor_and_term_seq.png" width="700"><br />
<p><B>Figure 2:</B> Sequences of the oligonucleotides used to make the Anchor and Terminator.<p><br />
</center><br />
<br><br />
<br />
<br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<br />
<div class="Outreach"><br />
<br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<br />
<br />
<h1>Anchor Variants and Binding Capacity</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<br />
<p><br />
<br />
<h3>Anchor Version #1</h3><br />
As seen in <B>Table 1</B>, we considered three main types of anchor systems. The simplest being a 5' biotinylated 20 nt ssDNA anchor. The product brochure for the paramagnetic streptavidin beads from NEB (Cat. # S1420S) claimed that the binding capacity of such an oligo is 500 pmol mg<sup>-1</sup> beads. We did not bother to confirm this because this anchor does not allow for ligation of incoming Bytes, since there is no complementary strand to which the incoming Byte may ligate. There is also no mechanism to release the construct from the bead, other than boiling the beads (which is not desireable). </p><br />
<br />
<p><br />
<h3>Anchor Version #2</h3><br />
A simple anchor system we had initially developed was one whereby we use 5'biotinylated forward primers (pAB/pBA forward sequencing primer) without the incorporation of deoxyuracils, and uracil containing universal reverse primers (pAB_R and pBA_R). PCR is conducted in the presence of the Byte you wish to make your Anchor and the result is amplified 5' biotinylated pAB/pBA insert (gene) with a 3' uracil containing end. The characteristic 12 base overhangs are generated by USER digestion, however since the biotinylated forward primers do not contain uracils, only the 3' end of the PCR product is acted upon by USER, thus only the 3' end of the "Byte" has an overhang. This was then directly bound to the beads. The one advantage of this system is the anchor itself is a Byte and contributes directly to the final construct size. However, it can only be released by NotI digestion (a consequence of the forward primer sequences used). Most importantly, it was found that this method of anchoring had terrible binding capacities, depending on the size of the anchor piece (2-8 pmol mg<sup>-1</sup> beads) due to the lack of a ssDNA spacer region between the 5' biotin and the dsDNA region, and that fact that binding capacity has an inverse relationship with anchor size.</p><br />
<br />
<p><br />
<h3>Anchor Version #3: The BioByte Anchor</h3><br />
The anchoring system we decided on was the one described above. It's small size and the presence of the ssDNA spacer region gives this anchor a high binding capacity of about 200 pmol mg<sup>-1</sup> beads. The presence of the dsDNA region allows Bytes to be ligated to the anchor and the incorporation of a deoxyuracil containing I-SceI site allows the construct to be released via the two methods described already(USER and I-SceI digestion).</p><br />
<br />
<p><br />
<h3>Binding Capacity Determination</h3><br />
Anchor system #2, whose protocol for binding capacity is not shown, was tested by binding a known high concentration of the biotinylated DNA to a known amount of beads. Binding capacity was determined by quantifying both the decrease in concentration of free anchor in solution after binding to the bead as well as measuring the amount of DNA is solution following enxymatic release from the bead, a direct measurement of the ng of DNA bound. This system of anchor, that is biotinylated gene-size dsDNA, had a paltry binding capacity of only 2-8 pmol mg<sup>-1</sup> beads.</p><br />
<br />
<p><br />
The bead binding capacity for the BioBytes anchor system was determined, see the protocol <a href="https://2009.igem.org/Team:Alberta/Project/BeadBindingCapacity">here</a>. The results are shown below.</p><br />
<br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/3/33/Alberta_iGEM-2009CapacityVSbeads.png" width="700"><br />
</center><br />
<br><br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/f/f4/Alberta_iGEM-2009_CapacityVSDNA.png" width="700"><br />
</center><br />
<br><br />
<br />
<br />
<p><br />
The graphs show a strong hyperbolic relationship of binding capacity between amount of beads used and a linear relationship to the concentration of DNA used. Thus the assay we have outlined in the protocols section is flawed. Having run the experiment multiple times and calculated similar binding capacities for the various volumes of beads we decided to just use the binding capacity for 40 uL of beads, since this is the amount we use for all our assemblies. The binding capacity for the BioBytes anchor system is 209 ± 20 pmol mg<sup>-1</sup> beads.</p><br />
<br />
<br />
</font><br />
<br />
</div></div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/Team:Alberta/DNAanchorTeam:Alberta/DNAanchor2009-10-22T03:42:18Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#e1e1e1; margin:0 5px;}<br />
.b2f {height:1px; background:#e1e1e1; margin:0 3px;}<br />
.b3f {height:1px; background:#e1e1e1; margin:0 2px;}<br />
.b4f {height:2px; background:#e1e1e1; margin:0 1px;}<br />
.content {background: #e1e1e1;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<br />
<h1>DNA Anchor/Terminator</h1><br />
<br />
<!-- <br />
<h3>Brick Creation</h3><br />
<br />
<p>The use of uracil-containing primers and USER<sup>TM</sup> enzyme mix provides an alternative method for creating 12 base sticky ends that is more versatile than conventional restriction endonucleases. The primers anneal to the A and B regions respectively, as well as ~5bp 3' into the cassette (to increase melting temperature). Bricks cloned into pAB and pBA can be PCR'd up with the universal uracil primers prA1u/prB1u (for pAB), or prA2u/prB2u(for pBA) and treated with USER<sup>TM</sup> mix. The uracil DNA glycosylase (UDG) present in the mix will cleave the uracil base off the DNA backbone. Endonuclease VIII will subsequently cleave the sugar-phosphate backbone at the apyrimidinic site generated by UDG, creating single stranded regions which can be purified away using PCR purification spin columns or gel purification. See <B>Figure 1</B>. </P><br />
<p> A protocol for generating Bytes can be found <a href="https://2009.igem.org/Team:Alberta/Project/ByteAmplification">here</a> and more information on Byte formation can be found <a href="https://2009.igem.org/Team:Alberta/Splasmids">here</a>.<br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/e/e0/UofA09_Bead_Overview_anchor2.png"><br />
<p><B>Figure 1</B>: Showing anchor and terminator fragments and effect of USER<sup>TM</sup> treatment. I SceI site and A ends are highlighted<p><br />
</center><br />
<br><br />
<br />
!--><br />
<br />
<h3>Anchoring System</h3><br />
<br />
<br />
<p>A vital component of the BioBytes method is the use of a biotinylated DNA Anchor in order to allow unidirectional assembly of the Bytes on paramagnetic beads by sequestering the 5' ends of Bytes, leaving only the 3' ends available to bind incomding Bytes. The Anchor itself has three vital components: A 5’ biotinylation, a double stranded DNA (dsDNA) portion that incorporates a release mechanism in order to liberate the construct from the beads, and A or B overhangs to allow Bytes to bind to the Anchor. Our team has considered a number anchor systems, each with their own set of advantages and disadvantages. The three main anchor systems we investigated are summarized below. See <B>Table 1</B>.</P><br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/f/f5/Anchorcomparison.png"><br />
<p><B>Table 1:</B> Overview of three different anchoring systems that were considered for BioBytes. See the section on <B>Anchor Variants and Binding Capacity</B> below for further information on these systems. 5' Biotin (BTN), single stranded DNA (ssDNA), nucleotide (nt), double stranded DNA (dsDNA).<p><br />
</center><br />
<br><br />
<br />
<p>The current BioBytes anchor system utilizes a 5’-biotin which anchors the construct to beads by binding non-covalently, but with great strength, to the covalently linked streptavidin on the surface of the paramagnetic beads. There is also a 5’-15 nucleotide spacer region of ssDNA which facilitates more efficient binding of the Anchor to the bead as the binding pocket of streptavidin is deep and thus a highly flexible ssDNA linker is recommended to allow the biotin to effectively bind into this deep pocket. A 21 bp double stranded portion of the Anchor contains the I-SceI recognition sequence, which when digested with I-SceI produces 4 base overhangs, but also includes four deoxyuracil residues. These uracils are excised by New England Biolab’s USER<sup>TM</sup> system to generate single nucleotide gaps in the top strand. USER<sup>TM</sup> digestion thus effectively destroys the Anchor and produces a 21 base 3’ overhang which becomes important for recircularization of the construct. Finally the Anchor contains the A or B 3’ overhangs complementary to those of the Bytes, allowing their binding to the Anchor. See <B>Figure 1</B>.</P><br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/e/e0/UofA09_Bead_Overview_anchor2.png"><br />
<p><B>Figure 1:</B> pAB and pBA multiple cloning sites with highlighted primers pAB_F/R and pBA_F/R annealing regions.<p><br />
</center><br />
<br><br />
<br />
<br />
<br />
<h3>Termination System</h3><br />
<br />
<p>Once the construct has been completed, i.e. the last Byte has been added, the construct may be released from the beads as is by a simple I-SceI digestion or a USER<sup>TM</sup> digestion, thus yielding a linear construct. If a circular construct, such as a plasmid, is desired then a final "Terminator" piece must be added. This piece is similar in construction to the Anchor, whereby there is a dsDNA I-SceI recognition sequence with four deoxyuracils incorporated into it on one strand, as well as an A or B 3' overhang. The Terminator binds to the last Byte and release is once again achieved by I-SceI digestion or USER<sup>TM</sup> digestion. In either case both the Anchor and Terminator develop sticky ends that are complementary to each other: 4 bases if I-SceI digestion is utilized, or 21 bases if USER is used. USER<sup>TM</sup> digestion is obviously preferred since 21 bp of interaction will form spontaneously and without ligation, and thus transformation of the construct can proceed immediately. See <B>Figure 1</B>.</P><br />
<br />
<h3>Anchor and Terminator Oligo Sequences</h3><br />
<p>The following sequences (<B>Figure 2</B>) are for the oligonucleotides one must order and anneal to generate the full set of Anchors and Terminators. The Anchor_A piece must be used in an Anchor that is meant to bind an AB Byte and Anchor_B for a BA Byte. Terminators containing the Term_A piece will bind BA Bytes, whereas those with a Term_B will bind AB Bytes; remembering that Terminators bind the the 3' end of Bytes and Anchors to the 5' end of Bytes. The Term_Comp and Anchor_Comp sequences are the complementary sequences that anneal to both the Term_A/B and Anchor_A/B pieces respectively to give the 21 bp dsDNA portion of both the Anchor and Terminator. Thus if you want to make an Anchor with an A overhang, you must anneal Anchor_A with Anchor_comp, etc.</p><br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/3/31/Anchor_and_term_seq.png" width="700"><br />
<p><B>Figure 2:</B> Sequences of the oligonucleotides used to make the Anchor and Terminator.<p><br />
</center><br />
<br><br />
<br />
<br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<br />
<div class="Outreach"><br />
<br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<br />
<br />
<h1>Anchor Variants and Binding Capacity</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<br />
<p><br />
<br />
<h3>Anchor Version #1</h3><br />
As seen in <B>Table 1</B>, we considered three main types of anchor systems. The simplest being a 5' biotinylated 20 nt ssDNA anchor. The product brochure for the paramagnetic streptavidin beads from NEB (Cat. # S1420S) claimed that the binding capacity of such an oligo is 500 pmol mg<sup>-1</sup> beads. We did not bother to confirm this because this anchor does not allow for ligation of incoming Bytes, since there is no complementary strand to which the incoming Byte may ligate. There is also no mechanism to release the construct from the bead, other than boiling the beads (which is not desireable). </p><br />
<br />
<p><br />
<h3>Anchor Version #2</h3><br />
A simple anchor system we had initially developed was one whereby we use 5'biotinylated forward primers (pAB/pBA forward sequencing primer) without the incorporation of deoxyuracils, and uracil containing universal reverse primers (pAB_R and pBA_R). PCR is conducted in the presence of the Byte you wish to make your Anchor and the result is amplified 5' biotinylated pAB/pBA insert (gene) with a 3' uracil containing end. The characteristic 12 base overhangs are generated by USER digestion, however since the biotinylated forward primers do not contain uracils, only the 3' end of the PCR product is acted upon by USER, thus only the 3' end of the "Byte" has an overhang. This was then directly bound to the beads. The one advantage of this system is the anchor itself is a Byte and contributes directly to the final construct size. However, it can only be released by NotI digestion (a consequence of the forward primer sequences used). Most importantly, it was found that this method of anchoring had terrible binding capacities, depending on the size of the anchor piece (2-8 pmol mg<sup>-1</sup> beads) due to the lack of a ssDNA spacer region between the 5' biotin and the dsDNA region, and that fact that binding capacity has an inverse relationship with anchor size.</p><br />
<br />
<p><br />
<h3>Anchor Version #3: The BioByte Anchor</h3><br />
The anchoring system we decided on was the one described above. It's small size and the presence of the ssDNA spacer region gives this anchor a high binding capacity of about 200 pmol mg<sup>-1</sup> beads. The presence of the dsDNA region allows Bytes to be ligated to the anchor and the incorporation of a deoxyuracil containing I-SceI site allows the construct to be released via the two methods described already(USER and I-SceI digestion).</p><br />
<br />
<p><br />
<h3>Binding Capacity Determination</h3><br />
Anchor system #2, whose protocol for binding capacity is not shown, was tested by binding a known high concentration of the biotinylated DNA to a known amount of beads. Binding capacity was determined by quantifying both the decrease in concentration of free anchor in solution after binding to the bead as well as measuring the amount of DNA is solution following enxymatic release from the bead, a direct measurement of the ng of DNA bound. This system of anchor, that is biotinylated gene-size dsDNA, had a paltry binding capacity of only 2-8 pmol mg<sup>-1</sup> beads.</p><br />
<br />
<p><br />
The bead binding capacity for the BioBytes anchor system was determined, see the protocol <a href="https://2009.igem.org/Team:Alberta/Project/BeadBindingCapacity">here</a>. The results are shown below.</p><br />
<br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/3/33/Alberta_iGEM-2009CapacityVSbeads.png" width="700"><br />
</center><br />
<br><br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/f/f4/Alberta_iGEM-2009_CapacityVSDNA.png" width="700"><br />
</center><br />
<br><br />
<br />
<br />
<p><br />
The graphs show a strong hyperbolic relationship of binding capacity to amount of beads used and a linear relationship to the concentration of DNA used. Thus the assay we have outlined in the protocols section is flawed. Having run the experiment multiple times and calculated similar binding capacities for the various volumes of beads we decided to just use the binding capacity for 40 uL of beads, since this is the amount we use for all our assemblies. The binding capacity for the BioBytes anchor system is 209 ± 20 pmol mg<sup>-1</sup> beads.</p><br />
<br />
<br />
</font><br />
<br />
</div></div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/Team:Alberta/DNAanchorTeam:Alberta/DNAanchor2009-10-22T03:39:44Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#e1e1e1; margin:0 5px;}<br />
.b2f {height:1px; background:#e1e1e1; margin:0 3px;}<br />
.b3f {height:1px; background:#e1e1e1; margin:0 2px;}<br />
.b4f {height:2px; background:#e1e1e1; margin:0 1px;}<br />
.content {background: #e1e1e1;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<br />
<h1>DNA Anchor/Terminator</h1><br />
<br />
<!-- <br />
<h3>Brick Creation</h3><br />
<br />
<p>The use of uracil-containing primers and USER<sup>TM</sup> enzyme mix provides an alternative method for creating 12 base sticky ends that is more versatile than conventional restriction endonucleases. The primers anneal to the A and B regions respectively, as well as ~5bp 3' into the cassette (to increase melting temperature). Bricks cloned into pAB and pBA can be PCR'd up with the universal uracil primers prA1u/prB1u (for pAB), or prA2u/prB2u(for pBA) and treated with USER<sup>TM</sup> mix. The uracil DNA glycosylase (UDG) present in the mix will cleave the uracil base off the DNA backbone. Endonuclease VIII will subsequently cleave the sugar-phosphate backbone at the apyrimidinic site generated by UDG, creating single stranded regions which can be purified away using PCR purification spin columns or gel purification. See <B>Figure 1</B>. </P><br />
<p> A protocol for generating Bytes can be found <a href="https://2009.igem.org/Team:Alberta/Project/ByteAmplification">here</a> and more information on Byte formation can be found <a href="https://2009.igem.org/Team:Alberta/Splasmids">here</a>.<br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/e/e0/UofA09_Bead_Overview_anchor2.png"><br />
<p><B>Figure 1</B>: Showing anchor and terminator fragments and effect of USER<sup>TM</sup> treatment. I SceI site and A ends are highlighted<p><br />
</center><br />
<br><br />
<br />
!--><br />
<br />
<h3>Anchoring System</h3><br />
<br />
<br />
<p>A vital component of the BioBytes method is the use of a biotinylated DNA Anchor in order to allow unidirectional assembly of the Bytes on paramagnetic beads by sequestering the 5' ends of Bytes, leaving only the 3' ends available to bind incomding Bytes. The Anchor itself has three vital components: A 5’ biotinylation, a double stranded DNA (dsDNA) portion that incorporates a release mechanism in order to liberate the construct from the beads, and A or B overhangs to allow Bytes to bind to the Anchor. Our team has considered a number anchor systems, each with their own set of advantages and disadvantages. The three main anchor systems we investigated are summarized below. See <B>Table 1</B>.</P><br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/f/f5/Anchorcomparison.png"><br />
<p><B>Table 1:</B> Overview of three different anchoring systems that were considered for BioBytes. See the section on <B>Anchor Variants and Binding Capacity</B> below for further information on these systems. 5' Biotin (BTN), single stranded DNA (ssDNA), nucleotide (nt), double stranded DNA (dsDNA).<p><br />
</center><br />
<br><br />
<br />
<p>The current BioBytes anchor system utilizes a 5’-biotin which anchors the construct to beads by binding non-covalently, but with great strength, to the covalently linked streptavidin on the surface of the paramagnetic beads. There is also a 5’-15 nucleotide spacer region of ssDNA which facilitates more efficient binding of the Anchor to the bead as the binding pocket of streptavidin is deep and thus a highly flexible ssDNA linker is recommended to allow the biotin to effectively bind into this deep pocket. A 21 bp double stranded portion of the Anchor contains the I-SceI recognition sequence, which when digested with I-SceI produces 4 base overhangs, but also includes four deoxyuracil residues. These uracils are excised by New England Biolab’s USER<sup>TM</sup> system to generate single nucleotide gaps in the top strand. USER<sup>TM</sup> digestion thus effectively destroys the Anchor and produces a 21 base 3’ overhang which becomes important for recircularization of the construct. Finally the Anchor contains the A or B 3’ overhangs complementary to those of the Bytes, allowing their binding to the Anchor. See <B>Figure 1</B>.</P><br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/e/e0/UofA09_Bead_Overview_anchor2.png"><br />
<p><B>Figure 1:</B> pAB and pBA multiple cloning sites with highlighted primers pAB_F/R and pBA_F/R annealing regions.<p><br />
</center><br />
<br><br />
<br />
<br />
<br />
<h3>Termination System</h3><br />
<br />
<p>Once the construct has been completed, i.e. the last Byte has been added, the construct may be released from the beads as is by a simple I-SceI digestion or a USER<sup>TM</sup> digestion, thus yielding a linear construct. If a circular construct, such as a plasmid, is desired then a final "Terminator" piece must be added. This piece is similar in construction to the Anchor, whereby there is a dsDNA I-SceI recognition sequence with four deoxyuracils incorporated into it on one strand, as well as an A or B 3' overhang. The Terminator binds to the last Byte and release is once again achieved by I-SceI digestion or USER<sup>TM</sup> digestion. In either case both the Anchor and Terminator develop sticky ends that are complementary to eachother: 4 bases if I-SceI digestion is utilized, or 21 bases if USER is used. USER<sup>TM</sup> digestion is obviously preferred since 21 bp of interaction will form spontaneously and without ligation, and thus transformation of the construct can proceed immediately. See <B>Figure 1</B>.</P><br />
<br />
<h3>Anchor and Terminator Oligo Sequences</h3><br />
<p>The following sequences (<B>Figure 2</B>) are for the oligonucleotides one must order and anneal to generate the full set of Anchors and Terminators. The Anchor_A piece must be used in an Anchor that is meant to bind an AB Byte and Anchor_B for a BA Byte. Terminators containing the Term_A piece will bind BA Bytes, whereas those with a Term_B will bind AB Bytes; remembering that Terminators bind the the 3' end of Bytes and Anchors to the 5' end of Bytes. The Term_Comp and Anchor_Comp sequences are the complementary sequences that anneal to both the Term_A/B and Anchor_A/B pieces respectively to give the 21 bp dsDNA portion of both the Anchor and Terminator. Thus if you want to make an Anchor with an A overhang, you must anneal Anchor_A with Anchor_comp, etc.</p><br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/3/31/Anchor_and_term_seq.png" width="700"><br />
<p><B>Figure 2:</B> Sequences of the oligonucleotides used to make the Anchor and Terminator.<p><br />
</center><br />
<br><br />
<br />
<br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<br />
<div class="Outreach"><br />
<br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<br />
<br />
<h1>Anchor Variants and Binding Capacity</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<br />
<p><br />
<br />
<h3>Anchor Version #1</h3><br />
As seen in <B>Table 1</B>, we considered three main types of anchor systems. The simplest being a 5' biotinylated 20 nt ssDNA anchor. The product brochure for the paramagnetic streptavidin beads from NEB (Cat. # S1420S) claimed that the binding capacity of such an oligo is 500 pmol mg<sup>-1</sup> beads. We did not bother to confirm this as this anchor does not allow for ligation of incoming Bytes to it, as there is no complementary strand for which the incoming Byte may ligate to. There is also no mechanism to release the construct from the bead, other than boiling the beads (which is not desireable). </p><br />
<br />
<p><br />
<h3>Anchor Version #2</h3><br />
A simple anchor system we had initially developed was one whereby we use 5'biotinylated forward primers (pAB/pBA forward sequencing primer) without the incorporation of deoxyuracils, and uracil containing universal reverse primers (pAB_R and pBA_R). PCR is conducted in the presence of the Byte you wish to make your Anchor and the result is amplified 5' biotinylated pAB/pBA insert (gene) with a 3' uracil containing end. The characteristic 12 base overhangs are generated by USER digestion, however since the biotinylated forward primers do not contain uracils, only the 3' end of the PCR product is acted upon by USER, thus only the 3' end of the "Byte" has an overhang. This was then directly bound to the beads. The one advantage of this system is the anchor itself is a Byte and contributes directly to the final construct size. However, it can only be released by NotI digestion (a consequence of the forward primer sequences used). Most importantly, it was found that this method of anchoring had terrible binding capacities, depending on the size of the anchor piece (2-8 pmol mg<sup>-1</sup> beads) due to the lack of a ssDNA spacer region between the 5' biotin and the dsDNA region, and that fact that binding capacity has an inverse relationship with anchor size.</p><br />
<br />
<p><br />
<h3>Anchor Version #3: The BioByte Anchor</h3><br />
The anchoring system we decided on was the one described above. It's small size and the presence of the ssDNA spacer region gives this anchor a high binding capacity of about 200 pmol mg<sup>-1</sup> beads. The presence of the dsDNA region allows Bytes to be ligated to the anchor and the incorporation of a deoxyuracil containing I-SceI site allows the construct to be released via the two methods described already(USER and I-SceI digestion).</p><br />
<br />
<p><br />
<h3>Binding Capacity Determination</h3><br />
Anchor system #2, whose protocol for binding capacity is not shown, was tested by binding a known high concentration of the biotinylated DNA to a known amount of beads. Binding capacity was determined by quantifying both the decrease in concentration of free anchor in solution after binding to the bead as well as measuring the amount of DNA is solution following enxymatic release from the bead, a direct measurement of the ng of DNA bound. This system of anchor, that is biotinylated gene-size dsDNA, had a paltry binding capacity of only 2-8 pmol mg<sup>-1</sup> beads.</p><br />
<br />
<p><br />
The bead binding capacity for the BioBytes anchor system was determined, see the protocol <a href="https://2009.igem.org/Team:Alberta/Project/BeadBindingCapacity">here</a>. The results are shown below.</p><br />
<br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/3/33/Alberta_iGEM-2009CapacityVSbeads.png" width="700"><br />
</center><br />
<br><br />
<br />
<br><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/f/f4/Alberta_iGEM-2009_CapacityVSDNA.png" width="700"><br />
</center><br />
<br><br />
<br />
<br />
<p><br />
The graphs show a strong hyperbolic relationship of binding capacity to amount of beads used and a linear relationship to the concentration of DNA used. Thus the assay we have outlined in the protocols section is flawed. Having run the experiment multiple times and calculated similar binding capacities for the various volumes of beads we decided to just use the binding capacity for 40 uL of beads, since this is the amount we use for all our assemblies. The binding capacity for the BioBytes anchor system is 209 ± 20 pmol mg<sup>-1</sup> beads.</p><br />
<br />
<br />
</font><br />
<br />
</div></div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/Team:Alberta/ByteCreationTeam:Alberta/ByteCreation2009-10-22T03:05:34Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#e1e1e1; margin:0 5px;}<br />
.b2f {height:1px; background:#e1e1e1; margin:0 3px;}<br />
.b3f {height:1px; background:#e1e1e1; margin:0 2px;}<br />
.b4f {height:2px; background:#e1e1e1; margin:0 1px;}<br />
.content {background: #e1e1e1;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<br />
<h1>Byte Creation</h1><br />
<br />
<h2>Standard Plasmids</h2><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<p>Each part to be assembled via the BioBytes method needs to have its ends altered to either the AB or the BA Byte type. To do this, unique plasmids were developed and named <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K187000">pAB</a> and <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K187001">pBA</a> (Figure 1). These plasmids were designed and constructed from pUC19 and contain the pMB1 high copy origin. Unique cassettes were designed containing PstI, XbaI, and primer annealing regions complementary to the ‘A’ and ‘B’ ends. The cassettes (Figure 2) were synthesized and inserted using two restriction sites (EcoRI and NsiI). This left an EcoRI site on the final plasmid as well a PstI scar site. The primer annealing regions in pAB are reverse to that in pBA so that compatible sticky ends can be produced in either plasmid. Genes can be inserted using XbaI and PstI (or an enzyme which produces a compatible sticky end). The plasmids were originally designed for a previous but now obsolete system, where the sticky ends were generated via nicking enzymes. </p><br />
<br />
<p><b>Figure 1.</b></p><br />
<center><br />
<table><br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/9/95/PAB.png" width="360"><br />
</td><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/e/e7/PBA.png" width="360"><br />
</td><br />
</tr><br />
</table><br />
</center><br />
<p><b>Figure 2.</b></p><br />
<center><img src="https://static.igem.org/mediawiki/2009/c/ce/UofA09_Bead_ABcassetteimg.png"><br />
<img src="https://static.igem.org/mediawiki/2009/9/96/UofA09_Bead_BAcassetteimg.png"><br />
<br />
</center><br />
<br />
<h2>How To Create a Byte</h2><br />
<br />
<br />
<p>In order to format a part as an AB or BA form Byte, the part first needs to be cloned in to pAB or pBA, respectively. This is done using a XbaI and PstI digest of both the insert and backbone, followed by ligation to place the part inside of the AB or BA cassette.</p><br />
<br />
<p>Once the part is cloned, universal PCR primers containing deoxyuridine residues are used to amplify the part. To create an AB Byte from pAB, the universal primers <a href="http://partsregistry.org/wiki/index.php/Part:BBa_K187365">pAB_F</a> and <a href="http://partsregistry.org/wiki/index.php/Part:BBa_K187366">pAB_R</a> are used. For the creation of a BA Byte from pBA, the primers <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K187367">pBA_F</a> and <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K187368">pBA_R</a> are used (Figure 3).<br />
<br />
<p><b>Figure 3.</b></p><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/f/ff/UofA09_Bead_universalprimers.png"><br />
</center><br />
<br />
After amplification, treatment with USER<sup>TM</sup> mix (available from New England Biolabs) creates a nucleotide gap at the position of the uracil by first excising the uracil base by Uracil DNA glycosylase and then cleaving the phosphodiester backbone at the apyrimidinic site via Endonuclease VIII. The resulting short oligonucleotides are then purified away from the PCR product to produce mature 12 base 3' overhangs of the AB or BA form Byte (Figure 4).</p><br />
<br />
<br />
<p><b>Figure 4.</b></p><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/c/c3/Alberta_byteconstruction.png" width="500"><br />
</center><br />
<br />
<P>The protocol for amplifying and digesting AB and BA Bytes can be found in our <a href="https://2009.igem.org/Team:Alberta/Protocols">lab section</a>.<br />
<br />
</P><br />
</div><br />
<br />
<br />
<br />
<br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
<br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/Team:Alberta/ByteCreationTeam:Alberta/ByteCreation2009-10-22T03:01:31Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#e1e1e1; margin:0 5px;}<br />
.b2f {height:1px; background:#e1e1e1; margin:0 3px;}<br />
.b3f {height:1px; background:#e1e1e1; margin:0 2px;}<br />
.b4f {height:2px; background:#e1e1e1; margin:0 1px;}<br />
.content {background: #e1e1e1;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<br />
<h1>Byte Creation</h1><br />
<br />
<h2>Standard Plasmids</h2><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<p>Each part to be assembled via the BioBytes method needs to have its ends altered to either the AB or the BA Byte type. To do this, unique plasmids were developed and named <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K187000">pAB</a> and <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K187001">pBA</a> (Figure 1). These plasmids were designed and constructed from pUC19 and contain the pMB1 high copy origin. Unique cassettes were designed containing PstI, XbaI, and primer annealing regions complementary to the ‘A’ and ‘B’ ends. The cassettes (Figure 2) were synthesized and inserted using two restriction sites (EcoRI and NsiI). This left an EcoRI site on the final plasmid as well a PstI scar site. The primer annealing regions in pAB are reverse to that in pBA so that compatible sticky ends can be produced in either plasmid. Genes can be inserted using XbaI and PstI (or an enzyme which produces a compatible sticky end). The plasmids were originally designed for a previous but now obsolete system, where the sticky ends were generated via nicking enzymes. </p><br />
<br />
<p><b>Figure 1.</b></p><br />
<center><br />
<table><br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/9/95/PAB.png" width="360"><br />
</td><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/e/e7/PBA.png" width="360"><br />
</td><br />
</tr><br />
</table><br />
</center><br />
<p><b>Figure 2.</b></p><br />
<center><img src="https://static.igem.org/mediawiki/2009/c/ce/UofA09_Bead_ABcassetteimg.png"><br />
<img src="https://static.igem.org/mediawiki/2009/9/96/UofA09_Bead_BAcassetteimg.png"><br />
<br />
</center><br />
<br />
<h2>How To Create a Byte</h2><br />
<br />
<br />
<p>In order to format a part as a AB or BA form Byte, the part first needs to be cloned in to pAB or pBA, respectively. This is done using a XbaI and PstI digest of both the insert and backbone, followed by ligation to place the part inside of the AB or BA cassette.</p><br />
<br />
<p>Once the part is cloned, universal PCR primers containing deoxyuridine residues are used to amplify the part. To create an AB Byte from pAB, the universal primers <a href="http://partsregistry.org/wiki/index.php/Part:BBa_K187365">pAB_F</a> and <a href="http://partsregistry.org/wiki/index.php/Part:BBa_K187366">pAB_R</a> are used. For the creation of a BA Byte from pBA, the primers <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K187367">pBA_F</a> and <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K187368">pBA_R</a> are used (Figure 3).<br />
<br />
<p><b>Figure 3.</b></p><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/f/ff/UofA09_Bead_universalprimers.png"><br />
</center><br />
<br />
After amplification, treatment with USER<sup>TM</sup> mix (available from New England Biolabs) creates a nucleotide gap at the position of the uracil by first excising the uracil base by Uracil DNA glycosylase and then cleaving the phosphodiester backbone at the apyrimidinic site via Endonuclease VIII. The resulting short oligonucleotides are then purified away from the PCR product to produce mature 12 base 3' overhangs of the AB or BA form Byte (Figure 4).</p><br />
<br />
<br />
<p><b>Figure 4.</b></p><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/c/c3/Alberta_byteconstruction.png" width="500"><br />
</center><br />
<br />
<P>The protocol for amplifying and digesting AB and BA Bytes can be found in our <a href="https://2009.igem.org/Team:Alberta/Protocols">lab section</a>.<br />
<br />
</P><br />
</div><br />
<br />
<br />
<br />
<br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
<br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/Team:Alberta/ByteCreationTeam:Alberta/ByteCreation2009-10-22T03:00:22Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#e1e1e1; margin:0 5px;}<br />
.b2f {height:1px; background:#e1e1e1; margin:0 3px;}<br />
.b3f {height:1px; background:#e1e1e1; margin:0 2px;}<br />
.b4f {height:2px; background:#e1e1e1; margin:0 1px;}<br />
.content {background: #e1e1e1;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<br />
<h1>Byte Creation</h1><br />
<br />
<h2>Standard Plasmids</h2><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<p>Each part to be assembled via the BioBytes method needs to have its ends altered to either the AB or the BA Byte type. To do this, unique plasmids were developed and named <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K187000">pAB</a> and <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K187001">pBA</a> (Figure 1). These plasmids were designed and constructed from pUC19 and contain the pMB1 high copy origin. Unique cassettes were designed containing PstI, XbaI, and primer annealing regions complementary to the ‘A’ and ‘B’ ends. The cassettes (Figure 2) were synthesized and inserted using two restriction sites (EcoRI and NsiI). This left an EcoRI site on the final plasmid as well a PstI scar site. The primer annealing regions in pAB are reverse to that in pBA so that compatible sticky ends can be produced in either plasmid. Genes can be inserted using XbaI and PstI (or an enzyme which produces a compatible sticky end). The plasmids were originally designed for a previous but now obsolete system, where the sticky ends were generated via nicking enzymes. </p><br />
<br />
<p><b>Figure 1.</b></p><br />
<center><br />
<table><br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/9/95/PAB.png" width="350"><br />
</td><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/e/e7/PBA.png" width="350"><br />
</td><br />
</tr><br />
</table><br />
</center><br />
<p><b>Figure 2.</b></p><br />
<center><img src="https://static.igem.org/mediawiki/2009/c/ce/UofA09_Bead_ABcassetteimg.png"><br />
<img src="https://static.igem.org/mediawiki/2009/9/96/UofA09_Bead_BAcassetteimg.png"><br />
<br />
</center><br />
<br />
<h2>How To Create a Byte</h2><br />
<br />
<br />
<p>In order to format a part as a AB or BA form Byte, the part first needs to be cloned in to pAB or pBA, respectively. This is done using a XbaI and PstI digest of both the insert and backbone, followed by ligation to place the part inside of the AB or BA cassette.</p><br />
<br />
<p>Once the part is cloned, universal PCR primers containing deoxyuridine residues are used to amplify the part. To create an AB Byte from pAB, the universal primers <a href="http://partsregistry.org/wiki/index.php/Part:BBa_K187365">pAB_F</a> and <a href="http://partsregistry.org/wiki/index.php/Part:BBa_K187366">pAB_R</a> are used. For the creation of a BA Byte from pBA, the primers <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K187367">pBA_F</a> and <a href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K187368">pBA_R</a> are used (Figure 3).<br />
<br />
<p><b>Figure 3.</b></p><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/f/ff/UofA09_Bead_universalprimers.png"><br />
</center><br />
<br />
After amplification, treatment with USER<sup>TM</sup> mix (available from New England Biolabs) creates a nucleotide gap at the position of the uracil by first excising the uracil base by Uracil DNA glycosylase and then cleaving the phosphodiester backbone at the apyrimidinic site via Endonuclease VIII. The resulting short oligonucleotides are then purified away from the PCR product to produce mature 12 base 3' overhangs of the AB or BA form Byte (Figure 4).</p><br />
<br />
<br />
<p><b>Figure 4.</b></p><br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/c/c3/Alberta_byteconstruction.png" width="500"><br />
</center><br />
<br />
<P>The protocol for amplifying and digesting AB and BA Bytes can be found in our <a href="https://2009.igem.org/Team:Alberta/Protocols">lab section</a>.<br />
<br />
</P><br />
</div><br />
<br />
<br />
<br />
<br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
<br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Nucleic_acid_purification_using_microfabricated_silicon_structuresTeam:Alberta/References/Publications/Nucleic acid purification using microfabricated silicon structures2009-10-21T22:53:57Z<p>Mitch p: </p>
<hr />
<div>Authors: Nathaniel C. Cady, Scott Stelickb and Carl A. Batt<br />
<br />
Biosensors and Bioelectronics, Volume 19, Issue 1, 30 October 2003, Pages 59-66 <br />
<br />
<br />
<b>Abstract:</b> A microfluidic device has been designed, fabricated and tested for its ability to purify bacteriophage lambda DNA and bacterial chromosomal DNA, a necessary prerequisite for its incorporation into a biosensor. This device consists of a microfabricated channel in which silica-coated pillars were etched to increase the surface area within the channel by 300–600%, when the etch depth is varied from 20 to 50 μm. DNA was selectively bound to these pillars in the presence of the chaotropic salt guanidinium isothiocyanate, followed by washing with ethanol and elution with low-ionic strength buffer. Positive pressure was used to move solutions through the device, removing the need for centrifugation steps. The binding capacity for DNA in the device was approximately 82 ng/cm2 and on average, 10% of the bound DNA could be purified and recovered in the first 50 μl of elution buffer. Additionally, the device removed approximately 87% of the protein from a cell lysate. Nucleic acids recovered from the device were efficiently amplified by the polymerase chain reaction suggesting the utility of these components in an integrated, DNA amplification-based biosensor. The miniaturized format of this purification device, along with its excellent purification characteristics make it an ideal component for nucleic acid-based biosensors, especially those in which nucleic acid amplification is a critical step.<br />
<br />
<br />
<br />
<br />
<br />
'''Link:''' [http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6TFC-48JK205-2&_user=1067472&_rdoc=1&_fmt=&_orig=search&_sort=d&_docanchor=&view=c&_searchStrId=1058543561&_rerunOrigin=scholar.google&_acct=C000051251&_version=1&_urlVersion=0&_userid=1067472&md5=18389e35f807a33ab1212516c6575b25 Science Direct]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Microfluidic_devices_fabricated_inpoly(dimethylsiloxane)_for_biological_studiesTeam:Alberta/References/Publications/Microfluidic devices fabricated inpoly(dimethylsiloxane) for biological studies2009-10-21T22:50:14Z<p>Mitch p: New page: Authors: Samuel K. Sia., George M. Whitesides Electrophoresis 2003, 24, 3563–3576 <b>Abstract:</b> This review describes microfluidic systems in poly(dimethylsiloxane) (PDMS) for biolo...</p>
<hr />
<div>Authors: Samuel K. Sia., George M. Whitesides<br />
<br />
Electrophoresis 2003, 24, 3563–3576<br />
<br />
<b>Abstract:</b> This review describes microfluidic systems in poly(dimethylsiloxane) (PDMS) for biological<br />
studies. Properties of PDMS that make it a suitable platform for miniaturized biological studies, techniques for fabricating PDMS microstructures, and methods for controlling fluid flow in microchannels are discussed. Biological procedures that have been miniaturized into PDMS-based microdevices include immunoassays, separation<br />
of proteins and DNA, sorting and manipulation of cells, studies of cells in microchannels exposed to laminar flows of fluids, and large-scale, combinatorial screening. The review emphasizes the advantages of miniaturization for biological analysis, such as efficiency of the device and special insights into cell biology.<br />
<br />
'''Link:''' [http://ftp.wtec.loyola.edu/robotics/us_workshop/June22/paper_review_whitesides2003microfluidics.pdf Electrophoresis]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Monolithic_MicrofabricatedValves_and_Pumps_by_MultilayerSoft_LithographyTeam:Alberta/References/Publications/Monolithic MicrofabricatedValves and Pumps by MultilayerSoft Lithography2009-10-21T22:45:36Z<p>Mitch p: New page: Authors: Marc A. Unger, Hou-Pu Chou, Todd Thorsen, Axel Scherer, Stephen R. Quake Science 7 April 2000: Vol. 288. no. 5463, pp. 113 - 116 <b>Abstract:</b> Soft lithography is an alterna...</p>
<hr />
<div>Authors: Marc A. Unger, Hou-Pu Chou, Todd Thorsen, Axel Scherer, Stephen R. Quake <br />
<br />
Science 7 April 2000: Vol. 288. no. 5463, pp. 113 - 116<br />
<br />
<b>Abstract:</b> Soft lithography is an alternative to silicon-based micromachining that uses replica molding of nontraditional elastomeric materials to fabricate stamps and microfluidic channels. We describe here an extension to the soft lithography paradigm, multilayer soft lithography, with which devices consisting of multiple layers may be fabricated from soft materials. We used this technique to build active microfluidic systems containing on-off valves, switching valves, and pumps entirely out of elastomer. The softness of these materials allows the device areas to be reduced by more than two orders of magnitude compared with silicon-based devices. The other advantages of soft lithography, such as rapid prototyping, ease of fabrication, and biocompatibility, are retained. <br />
<br />
'''Link:''' [http://www.sciencemag.org/cgi/content/abstract/288/5463/113 Science]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/On-chip_magnetic_bead_microarray_using_hydrodynamic_focusing_in_apassive_magnetic_separatorTeam:Alberta/References/Publications/On-chip magnetic bead microarray using hydrodynamic focusing in apassive magnetic separator2009-10-21T22:41:02Z<p>Mitch p: New page: Authors: Smistrup K, Kjeldsen BG, Reimers JL, Dufva M, Petersen J, Hansen MF Lab on a Chip. 2005 Nov;5(11):1315-9 <b>Abstract:</b> Implementing DNA and protein microarrays into lab-on-a-...</p>
<hr />
<div>Authors: Smistrup K, Kjeldsen BG, Reimers JL, Dufva M, Petersen J, Hansen MF<br />
<br />
Lab on a Chip. 2005 Nov;5(11):1315-9<br />
<br />
<b>Abstract:</b> Implementing DNA and protein microarrays into lab-on-a-chip systems can be problematic since these are sensitive to heat and strong chemicals. Here, we describe the functionalization of a microchannel with two types of magnetic beads using hydrodynamic focusing combined with a passive magnetic separator with arrays of soft magnetic elements. The soft magnetic elements placed on both sides of the channel are magnetized by a relatively weak applied external magnetic field (21 mT) and provide magnetic field gradients attracting magnetic beads. Flows with two differently functionalized magnetic beads and a separating barrier flow are introduced simultaneously at the two channel sides and the centre of the microfluidic channel, respectively. On-chip experiments with fluorescence labeled beads demonstrate that the two types of beads are captured at each of the channel sidewalls. On-chip hybridization experiments show that the microfluidic systems can be functionalized with two sets of beads carrying different probes that selectively recognize a single base pair mismatch in target DNA. By switching the places of the two types of beads it is shown that the microsystem can be cleaned and functionalized repeatedly with different beads with no cross-talk between experiments.<br />
<br />
'''Link:''' [http://www.ncbi.nlm.nih.gov/pubmed/16234958 PubMed]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/PCR_microfluidic_devices_for_DNA_amplificationTeam:Alberta/References/Publications/PCR microfluidic devices for DNA amplification2009-10-21T22:37:42Z<p>Mitch p: New page: Authors: Zhang C, Xu J, Ma W, Zheng W. Biotechnology Advances, 2006, May-Jun; 24(3):243-84 <b>Abstract:</b> The miniaturization of biological and chemical analytical devices by micro-ele...</p>
<hr />
<div>Authors: Zhang C, Xu J, Ma W, Zheng W.<br />
<br />
Biotechnology Advances, 2006, May-Jun; 24(3):243-84<br />
<br />
<b>Abstract:</b> The miniaturization of biological and chemical analytical devices by micro-electro-mechanical-systems (MEMS) technology has posed a vital influence on such fields as medical diagnostics, microbial detection and other bio-analysis. Among many miniaturized analytical devices, the polymerase chain reaction (PCR) microchip/microdevices are studied extensively, and thus great progress has been made on aspects of on-chip micromachining (fabrication, bonding and sealing), choice of substrate materials, surface chemistry and architecture of reaction vessel, handling of necessary sample fluid, controlling of three or two-step temperature thermocycling, detection of amplified nucleic acid products, integration with other analytical functional units such as sample preparation, capillary electrophoresis (CE), DNA microarray hybridization, etc. However, little has been done on the review of above-mentioned facets of the PCR microchips/microdevices including the two formats of flow-through and stationary chamber in spite of several earlier reviews [Zorbas, H. Miniature continuous-flow polymerase chain reaction: a breakthrough? Angew Chem Int Ed 1999; 38 (8):1055-1058; Krishnan, M., Namasivayam, V., Lin, R., Pal, R., Burns, M.A. Microfabricated reaction and separation systems. Curr Opin Biotechnol 2001; 12:92-98; Schneegabeta, I., Köhler, J.M. Flow-through polymerase chain reactions in chip themocyclers. Rev Mol Biotechnol 2001; 82:101-121; deMello, A.J. DNA amplification: does 'small' really mean 'efficient'? Lab Chip 2001; 1: 24N-29N; Mariella, Jr. R. MEMS for bio-assays. Biomed Microdevices 2002; 4 (2):77-87; deMello AJ. Microfluidics: DNA amplification moves on. Nature 2003; 422:28-29; Kricka, L.J., Wilding, P. Microchip PCR. Anal BioAnal Chem 2003; 377:820-825]. In this review, we survey the advances of the above aspects among the PCR microfluidic devices in detail. Finally, we also illuminate the potential and practical applications of PCR microfluidics to some fields such as microbial detection and disease diagnosis, based on the DNA/RNA templates used in PCR microfluidics. It is noted, especially, that this review is to help a novice in the field of on-chip PCR amplification to more easily find the original papers, because this review covers almost all of the papers related to on-chip PCR microfluidics.<br />
<br />
'''Link:''' [http://www.ncbi.nlm.nih.gov/pubmed/16326063 PubMed]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Microfluidic_PicoArray_synthesis_ofoligodeoxynucleotides_and_simultaneous_assembling_of_multiple_DNA_sequencesTeam:Alberta/References/Publications/Microfluidic PicoArray synthesis ofoligodeoxynucleotides and simultaneous assembling of multiple DNA sequences2009-10-21T22:33:15Z<p>Mitch p: New page: Authors: Xiaochuan Zhou, Shiying Cai, Ailing Hong, Qimin You, Peilin Yu, Nijing Sheng, Onnop Srivannavit, Seema Muranjan, Jean Marie Rouillard, Yongmei Xia, Xiaolin Zhang, Qin Xiang, Renuk...</p>
<hr />
<div>Authors: Xiaochuan Zhou, Shiying Cai, Ailing Hong, Qimin You, Peilin Yu, Nijing Sheng, Onnop Srivannavit, Seema Muranjan, Jean Marie Rouillard, Yongmei Xia, Xiaolin Zhang, Qin Xiang, Renuka Ganesh, Qi Zhu, Anna Matejko, Erdogan Gulari and Xiaolian Gao<br />
<br />
Nucleic Acids Research 2004 32(18):5409-5417<br />
<br />
<b>Abstract:</b> Large DNA constructs of arbitrary sequences can currently be assembled with relative ease by joining short synthetic oligodeoxynucleotides (oligonucleotides). The ability to mass produce these synthetic genes readily will have a significant impact on research in biology and medicine. Presently, high-throughput gene synthesis is unlikely, due to the limits of oligonucleotide synthesis. We describe a microfluidic PicoArray method for the simultaneous synthesis and purification of oligonucleotides that are designed for multiplex gene synthesis. Given the demand for highly pure oligonucleotides in gene synthesis processes, we used a model to improve key reaction steps in DNA synthesis. The oligonucleotides obtained were successfully used in ligation under thermal cycling conditions to generate DNA constructs of several hundreds of base pairs. Protein expression using the gene thus synthesized was demonstrated. We used a DNA assembly strategy, i.e. ligation followed by fusion PCR, and achieved effective assembling of up to 10 kb DNA constructs. These results illustrate the potential of microfluidics-based ultra-fast oligonucleotide parallel synthesis as an enabling tool for modern synthetic biology applications, such as the construction of genome-scale molecular clones and cell-free large scale protein expression. <br />
<br />
'''Link:''' [http://nar.oxfordjournals.org/cgi/content/abstract/32/18/5409 Oxford Journals]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Lab-on-a-chip:_applications_in_proteomicsTeam:Alberta/References/Publications/Lab-on-a-chip: applications in proteomics2009-10-21T22:28:29Z<p>Mitch p: New page: Authors: Stephane Mouradian Current Opinion in Chemical Biology, Volume 6, Issue 1, 1 February 2002, Pages 51-56 <b>Abstract:</b> Recent advances in chip-based separation of proteins p...</p>
<hr />
<div>Authors: Stephane Mouradian<br />
<br />
Current Opinion in Chemical Biology, Volume 6, Issue 1, 1 February 2002, Pages 51-56 <br />
<br />
<b>Abstract:</b> Recent advances in chip-based separation of proteins provide methods that are faster and more convenient than conventional gel electrophoresis. Rapid and automated protein sizing on a chip is at the commercial stage and first attempts have been made to perform two-dimensional separation on a chip. Numerous designs have been described to interface a microfluidic chip to a mass spectrometer. Impressive integration efforts are demonstrated by the ability to perform on-chip trypsin digestion, separation and injection into a mass spectrometer with a single device.<br />
<br />
'''Link:''' [http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6VRX-451DFYG-B&_user=1067472&_rdoc=1&_fmt=&_orig=search&_sort=d&_docanchor=&view=c&_searchStrId=1058521203&_rerunOrigin=google&_acct=C000051251&_version=1&_urlVersion=0&_userid=1067472&md5=05dd016be236adb88cd9d1990cf78ca6 Science Direct]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Accurate_multiplex_gene_synthesisfrom_programmable_DNA_microchipsTeam:Alberta/References/Publications/Accurate multiplex gene synthesisfrom programmable DNA microchips2009-10-21T22:23:49Z<p>Mitch p: New page: Authors: Tian, JD., Gong, H., Sheng, NJ., Zhou, XC., Gulari, E., Gao, XL., Church, G Nature 432(7020): 1050-1054. Dec 2004 <b>Abstract:</b> Testing the many hypotheses from genomics and ...</p>
<hr />
<div>Authors: Tian, JD., Gong, H., Sheng, NJ., Zhou, XC., Gulari, E., Gao, XL., Church, G<br />
<br />
Nature 432(7020): 1050-1054. Dec 2004<br />
<br />
<b>Abstract:</b> Testing the many hypotheses from genomics and systems biology experiments demands accurate and cost-effective gene and genome synthesis. Here we describe a microchip-based technology for multiplex gene synthesis. Pools of thousands of 'construction' oligonucleotides and tagged complementary 'selection' oligonucleotides are synthesized on photo-programmable microfluidic chips(1), released, amplified and selected by hybridization to reduce synthesis errors ninefold. A one-step polymerase assembly multiplexing reaction assembles these into multiple genes. This technology enabled us to synthesize all 21 genes that encode the proteins of the Escherichia coli 30S ribosomal subunit, and to optimize their translation efficiency in vitro through alteration of codon bias. This is a significant step towards the synthesis of ribosomes in vitro and should have utility for synthetic biology in general.<br />
<br />
'''Link:''' [http://deepblue.lib.umich.edu/handle/2027.42/62677 Deep Blue]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/The_Digital_Revolution:_A_New_Paradigm_forMicrofluidicsTeam:Alberta/References/Publications/The Digital Revolution: A New Paradigm forMicrofluidics2009-10-21T22:19:01Z<p>Mitch p: New page: Authors: Mohamed Abdelgawad, and Aaron R. Wheeler Advanced Materials 2009, 21, 920–925 <b>Abstract:</b> The digital revolution has come to microfluidics. In digital microfluidics(DMF),...</p>
<hr />
<div>Authors: Mohamed Abdelgawad, and Aaron R. Wheeler<br />
<br />
Advanced Materials 2009, 21, 920–925<br />
<br />
<b>Abstract:</b> The digital revolution has come to microfluidics. In digital microfluidics(DMF), discrete droplets are manipulated by applying electrical fields to an array of electrodes. In contrast to microchannels, in DMF each sample and reagent is individually addressable, which facilitates exquisite control over chemical reactions. Here, we review the state-of-the-art in DMF, with a discussion of device formats, actuation physics, and biological and nonbiological applications. Along the way, we identify the key players in the field, and speculate on the advances and challenges that lie ahead. As with other fronts in the digital revolution, there have been and will be unexpected developments as DMF matures, but we posit that the future is bright for this<br />
promising technology.<br />
<br />
'''Link:''' [http://www.chem.utoronto.ca/staff/WHEELER/Papers/DigRevolution.pdf University of Toronto]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/A_microfluidic_mammalian_cell_sorter_based_onfluorescence_detectionTeam:Alberta/References/Publications/A microfluidic mammalian cell sorter based onfluorescence detection2009-10-21T22:13:24Z<p>Mitch p: </p>
<hr />
<div>Authors: V. Studer, , R. Jameson, E. Pellereau, A. Pépin and Y. Chen<br />
<br />
Microelectronic Engineering, Volumes 73-74, June 2004, Pages 852-857<br />
<br />
<b>Abstract:</b> We report on the development of microfluidic devices for single mammalian cell sorting and manipulation. These microfluidic devices are fabricated out of polydimethylsiloxane (PDMS) by multilayer soft lithography. They consist of several active units (mixer, pumps) pneumatically actuated by monolithic soft microvalves. Using this fabrication method we were able to develop a microfluidic device for the fast sorting of 10 μm diameter fluorescently tagged rare objects (mammalian cells or beads) sparsely distributed within a concentrated solution of non-tagged objects. We show that once sorted, these objects can be individually recovered in a small volume (nanolitre range) for further biochemical assays such as cell lysis, mRNA extraction and polymerase chain reaction.<br />
<br />
'''Link:''' [http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6V0W-4C62R1D-3&_user=1067472&_rdoc=1&_fmt=&_orig=search&_sort=d&_docanchor=&view=c&_searchStrId=1058511935&_rerunOrigin=scholar.google&_acct=C000051251&_version=1&_urlVersion=0&_userid=1067472&md5=b6052cab819e7688d099b5b53e49b586 Science Direct]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/A_microfluidic_mammalian_cell_sorter_based_onfluorescence_detectionTeam:Alberta/References/Publications/A microfluidic mammalian cell sorter based onfluorescence detection2009-10-21T22:11:48Z<p>Mitch p: New page: Authors: V. Studer, , R. Jameson, E. Pellereau, A. Pépin and Y. Chen Microelectronic Engineering, Volumes 73-74, June 2004, Pages 852-857 <b>Abstract:</b> We report on the development ...</p>
<hr />
<div>Authors: V. Studer, , R. Jameson, E. Pellereau, A. Pépin and Y. Chen<br />
<br />
Microelectronic Engineering, Volumes 73-74, June 2004, Pages 852-857<br />
<br />
<b>Abstract:</b> We report on the development of microfluidic devices for single mammalian cell sorting and manipulation. These microfluidic devices are fabricated out of polydimethylsiloxane (PDMS) by multilayer soft lithography. They consist of several active units (mixer, pumps) pneumatically actuated by monolithic soft microvalves. Using this fabrication method we were able to develop a microfluidic device for the fast sorting of 10 μm diameter fluorescently tagged rare objects (mammalian cells or beads) sparsely distributed within a concentrated solution of non-tagged objects. We show that once sorted, these objects can be individually recovered in a small volume (nanolitre range) for further biochemical assays such as cell lysis, mRNA extraction and polymerase chain reaction.<br />
<br />
'''Link:''' [ http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6V0W-4C62R1D-3&_user=1067472&_rdoc=1&_fmt=&_orig=search&_sort=d&_docanchor=&view=c&_searchStrId=1058511935&_rerunOrigin=scholar.google&_acct=C000051251&_version=1&_urlVersion=0&_userid=1067472&md5=b6052cab819e7688d099b5b53e49b586 Science Direct]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Small_volume_PCR_in_PDMS_biochips_with_integrated_fluidcontrol_and_vapour_barrierTeam:Alberta/References/Publications/Small volume PCR in PDMS biochips with integrated fluidcontrol and vapour barrier2009-10-21T22:08:50Z<p>Mitch p: New page: Authors: A. Ranjit Prakasha, S. Adamiab, c, V. Siebena, P. Pilarskia, L.M. Pilarskib, c and C.J. Backhouse Sensors and Actuators B: Chemical, Volume 113, Issue 1, 17 January 2006, Pages 3...</p>
<hr />
<div>Authors: A. Ranjit Prakasha, S. Adamiab, c, V. Siebena, P. Pilarskia, L.M. Pilarskib, c and C.J. Backhouse<br />
<br />
Sensors and Actuators B: Chemical, Volume 113, Issue 1, 17 January 2006, Pages 398-409 <br />
<br />
<b>Abstract:</b> In this paper we demonstrate a new method for microfabricating PDMS devices that controls vapour diffusion, thereby reducing water loss at elevated temperatures and greatly increasing the reliability of the PCR. In the past, the vapour and liquid diffusion properties of the PDMS material in microfluidic devices have impaired performance. We show that this water loss is primarily due to vapour diffusion from the PDMS biochip and by implanting a polyethylene vapour barrier layer in the PDMS, the overall fluid loss was almost eliminated (reduced by a factor of 3). We have also developed a procedure to ensure irreversible bonding between the PDMS and the implant. With this improved microfabrication method we demonstrate the feasibility and advantages of performing small volume PCR genetic amplification (i.e. with less than 2 μl of PCR sample) within a PDMS–glass hybrid biochip. Diaphragm pumps and pinch-off valves were integrated in the system and these enabled fluid retention during the amplification stage and will facilitate higher levels of on-chip automation.<br />
<br />
'''Link:''' [http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6THH-4G27SNR-2&_user=1067472&_rdoc=1&_fmt=&_orig=search&_sort=d&_docanchor=&view=c&_searchStrId=1058507098&_rerunOrigin=scholar.google&_acct=C000051251&_version=1&_urlVersion=0&_userid=1067472&md5=60821287bfa98f1925cb21bc421901f8 Science Direct]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Microfluidic_chip:_Next_generation_platform_for_systems_biologyTeam:Alberta/References/Publications/Microfluidic chip: Next generation platform for systems biology2009-10-21T22:05:06Z<p>Mitch p: </p>
<hr />
<div>Authors:Feng X, Du W, Luo Q, Liu BF<br />
<br />
Analytica Chimica Acta. 2009 Sep 14;650(1):83-97<br />
<br />
<b>Abstract:</b> Systems biology advocates the understanding of biology at the systems-level, which requires massive information of correlations among individual components in complex biological systems. Such comprehensive investigation entails the use of high-throughput analytical tools. Microfluidic technology holds high promise to facilitate the progress of biology by enabling miniaturization and upgrading of current biological research tools due to its advantages such as low sample consumption, reduced analysis time, high-throughput and compatible sizes with most biological samples. In this article, we documented the recent applications of microfluidic chips in biological researches at the molecular level, cellular level and organism level, serving the purpose for systems-level understanding of biology.<br />
<br />
'''Link:''' [http://www.ncbi.nlm.nih.gov/pubmed/19720178 PubMed]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/On-chip_transformation_of_bacteriaTeam:Alberta/References/Publications/On-chip transformation of bacteria2009-10-21T22:04:34Z<p>Mitch p: </p>
<hr />
<div>Authors: Kuniaki Nagamine, Shiho Onodera, Yu-suke Torisawa, Tomoyuki Yasukawa, Hitoshi Shiku, and Tomokazu Matsue<br />
<br />
Analytical Chemistry, 2005, 77 (13), pp 4278–4281<br />
<br />
<b>Abstract:</b> On-chip transformation of Escherichia coli cells was accomplished for the first time using a microbial array chip. The continuous E. coli transformation procedures were performed on a chip in which the microcompartment was composed of PDMS microfluidic channels and a silicon substrate predeposited with different plasmid DNAs. The PDMS microfluidic device enabled the parallel transformation of E. coli cells with various plasmid DNAs by separating each transformation area. The phenotypic differences reflecting different plasmid DNAs were identified by various approaches such as colorimetry, fluorometry, and electrochemical methods. This microbial array chip could become a versatile tool for many cell biological applications. <br />
<br />
'''Link:''' [http://pubs.acs.org/doi/abs/10.1021/ac048278n ACS Publications]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/On-chip_transformation_of_bacteriaTeam:Alberta/References/Publications/On-chip transformation of bacteria2009-10-21T22:04:05Z<p>Mitch p: New page: Authors: Kuniaki Nagamine, Shiho Onodera, Yu-suke Torisawa, Tomoyuki Yasukawa, Hitoshi Shiku, and Tomokazu Matsue Analytical Chemistry, 2005, 77 (13), pp 4278–4281 <b>Abstract:</b> On-...</p>
<hr />
<div>Authors: Kuniaki Nagamine, Shiho Onodera, Yu-suke Torisawa, Tomoyuki Yasukawa, Hitoshi Shiku, and Tomokazu Matsue<br />
<br />
Analytical Chemistry, 2005, 77 (13), pp 4278–4281<br />
<br />
<b>Abstract:</b> On-chip transformation of Escherichia coli cells was accomplished for the first time using a microbial array chip. The continuous E. coli transformation procedures were performed on a chip in which the microcompartment was composed of PDMS microfluidic channels and a silicon substrate predeposited with different plasmid DNAs. The PDMS microfluidic device enabled the parallel transformation of E. coli cells with various plasmid DNAs by separating each transformation area. The phenotypic differences reflecting different plasmid DNAs were identified by various approaches such as colorimetry, fluorometry, and electrochemical methods. This microbial array chip could become a versatile tool for many cell biological applications. <br />
<br />
'''Link:'''[http://pubs.acs.org/doi/abs/10.1021/ac048278n ACS Publications]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Microfluidic_chip:_Next_generation_platform_for_systems_biologyTeam:Alberta/References/Publications/Microfluidic chip: Next generation platform for systems biology2009-10-21T22:00:26Z<p>Mitch p: </p>
<hr />
<div>Authors:Feng X, Du W, Luo Q, Liu BF<br />
<br />
Analytica Chimica Acta. 2009 Sep 14;650(1):83-97<br />
<br />
<b>Abstract:</b> Systems biology advocates the understanding of biology at the systems-level, which requires massive information of correlations among individual components in complex biological systems. Such comprehensive investigation entails the use of high-throughput analytical tools. Microfluidic technology holds high promise to facilitate the progress of biology by enabling miniaturization and upgrading of current biological research tools due to its advantages such as low sample consumption, reduced analysis time, high-throughput and compatible sizes with most biological samples. In this article, we documented the recent applications of microfluidic chips in biological researches at the molecular level, cellular level and organism level, serving the purpose for systems-level understanding of biology.<br />
<br />
'''Link:'''[http://www.ncbi.nlm.nih.gov/pubmed/19720178 PubMed]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/An_integrated_microfluidic_biochemical_detection_system_forprotein_analysis_with_magnetic_bead-based_samplingcapabilitiesTeam:Alberta/References/Publications/An integrated microfluidic biochemical detection system forprotein analysis with magnetic bead-based samplingcapabilities2009-10-21T21:59:47Z<p>Mitch p: </p>
<hr />
<div>Authors: Choi JW, Oh KW, Thomas JH, Heineman WR, Halsall HB, Nevin JH, Helmicki AJ, Henderson HT, Ahn CH. <br />
<br />
Lab on a Chip 2002. Feb; 2(1): 27-30<br />
<br />
<br />
<b>Abstract:</b> This paper presents the development and characterization of an integrated microfluidic biochemical detection system for fast and low-volume immunoassays using magnetic beads, which are used as both immobilization surfaces and bio-molecule carriers. Microfluidic components have been developed and integrated to construct a microfluidic biochemical detection system. Magnetic bead-based immunoassay, as a typical example of biochemical detection and analysis, has been successfully performed on the integrated microfluidic biochemical analysis system that includes a surface-mounted biofilter and electrochemical sensor on a glass microfluidic motherboard. Total time required for an immunoassay was less than 20 min including sample incubation time, and sample volume wasted was less than 50 microl during five repeated assays. Fast and low-volume biochemical analysis has been successfully achieved with the developed biofilter and immunosensor, which is integrated to the microfluidic system. Such a magnetic bead-based biochemical detection system, described in this paper, can be applied to protein analysis systems.<br />
<br />
<br />
<br />
<br />
<br />
'''Link:''' [http://www.ncbi.nlm.nih.gov/pubmed/15100857 PubMed]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Microfluidic_chip:_Next_generation_platform_for_systems_biologyTeam:Alberta/References/Publications/Microfluidic chip: Next generation platform for systems biology2009-10-21T21:58:23Z<p>Mitch p: New page: Authors:Feng X, Du W, Luo Q, Liu BF Anal Chim Acta. 2009 Sep 14;650(1):83-97 <b>Abstract:</b> Systems biology advocates the understanding of biology at the systems-level, which requires ...</p>
<hr />
<div>Authors:Feng X, Du W, Luo Q, Liu BF<br />
<br />
Anal Chim Acta. 2009 Sep 14;650(1):83-97<br />
<br />
<b>Abstract:</b> Systems biology advocates the understanding of biology at the systems-level, which requires massive information of correlations among individual components in complex biological systems. Such comprehensive investigation entails the use of high-throughput analytical tools. Microfluidic technology holds high promise to facilitate the progress of biology by enabling miniaturization and upgrading of current biological research tools due to its advantages such as low sample consumption, reduced analysis time, high-throughput and compatible sizes with most biological samples. In this article, we documented the recent applications of microfluidic chips in biological researches at the molecular level, cellular level and organism level, serving the purpose for systems-level understanding of biology.<br />
<br />
'''Link:'''[http://www.ncbi.nlm.nih.gov/pubmed/19720178 PubMed]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Electroporation_of_cells_in_microfluidic_devices:_a_reviewTeam:Alberta/References/Publications/Electroporation of cells in microfluidic devices: a review2009-10-21T17:47:50Z<p>Mitch p: </p>
<hr />
<div>Authors: M. B. Fox, D. C. Esveld, A. Valero, R. Luttge, H. C. Mastwijk, P. V. Bartels, A. van den Berg and R. M. Boom<br />
<br />
Analytical and Bioanalytical Chemistry, Volume 385, Number 3 / June, 2006 pp.474-485<br />
<br />
<b>Abstract:</b> In recent years, several publications on microfluidic devices have focused on the process of electroporation, which results in the poration of the biological cell membrane. The devices involved are designed for cell analysis, transfection or pasteurization. The high electric field strengths needed are induced by placing the electrodes in close proximity or by creating a constriction between the electrodes, which focuses the electric field. Detection is usually achieved through fluorescent labeling or by measuring impedance. So far, most of these devices have only concerned themselves solely with the electroporation process, but integration with separation and detection processes is expected in the near future. In particular, single-cell content analysis is expected to add further value to the concept of the microfluidic chip. Furthermore, if advanced pulse schemes are employed, such microdevices can also enhance research into intracellular electroporation.<br />
<br />
'''Link:''' [http://www.springerlink.com/content/f503582767v045t4/ Springer Link]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Electroporation_of_cells_in_microfluidic_devices:_a_reviewTeam:Alberta/References/Publications/Electroporation of cells in microfluidic devices: a review2009-10-21T17:46:27Z<p>Mitch p: New page: Authors: M. B. Fox, D. C. Esveld, A. Valero, R. Luttge, H. C. Mastwijk, P. V. Bartels, A. van den Berg and R. M. Boom Analytical and Bioanalytical Chemistry, Volume 385, Number 3 / June, ...</p>
<hr />
<div>Authors: M. B. Fox, D. C. Esveld, A. Valero, R. Luttge, H. C. Mastwijk, P. V. Bartels, A. van den Berg and R. M. Boom<br />
<br />
Analytical and Bioanalytical Chemistry, Volume 385, Number 3 / June, 2006 pp.474-485<br />
<br />
<b>Abstract:</b> In recent years, several publications on microfluidic devices have focused on the process of electroporation, which results in the poration of the biological cell membrane. The devices involved are designed for cell analysis, transfection or pasteurization. The high electric field strengths needed are induced by placing the electrodes in close proximity or by creating a constriction between the electrodes, which focuses the electric field. Detection is usually achieved through fluorescent labeling or by measuring impedance. So far, most of these devices have only concerned themselves solely with the electroporation process, but integration with separation and detection processes is expected in the near future. In particular, single-cell content analysis is expected to add further value to the concept of the microfluidic chip. Furthermore, if advanced pulse schemes are employed, such microdevices can also enhance research into intracellular electroporation.</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/On-chip_oligonucleotide_ligation_assay_using_one-dimensional_micro%EF%AC%82uidic_beads_array_for_the_detection_of_low-abundant_DNA_point_mutationsTeam:Alberta/References/Publications/On-chip oligonucleotide ligation assay using one-dimensional microfluidic beads array for the detection of low-abundant DNA point mutations2009-10-21T17:26:21Z<p>Mitch p: </p>
<hr />
<div>Authors: He Zhanga, Xiaohai Yanga, Kemin Wanga, Weihong Tana, Huimin Lib, Xinbing Zuoa and Jianhui Wena<br />
<br />
<br />
Biosensors and Bioelectronics Volume 23, Issue 7, 28 February 2008, Pages 945-951 <br />
<br />
<br />
<b>Abstract:</b> The detection of low-abundant DNA point mutations is very important for the early prediction of cancer, diagnostics of disease and clinical prognosis. In this paper, an on-chip oligonucleotide ligation approach that arrayed a series of functionalized beads in a single microfluidic channel was described for detection of low-abundant point mutations in p53 gene. This gene carried the point mutation with high diagnostic value for assessment of tumor progression and resectional borders. This work extended our prior efforts using one-dimensional (1-D) microfluidic beads array for protein and nucleic acid molecular profiling, and displayed high discrimination sensitivity to mutations detection due to the enhanced mass transport capability caused by microfluidic addressing format of beads array. As a demonstration, it was found that the on-chip beads ligation held high mutation discrimination sensitivity in 1 pM quantities at a SNR (signal-to-noise ratio) >2 using synthesized DNA oligonucleotides in accordance with target fragment. The RT-PCR products of tumor cell line A549, CNE2 and SKBr-3 were further examined to distinguish the point mutation at codon 175 of p53 gene. This approach was capable of detecting a point mutation in a p53 oncogene at a level of 1 mutant in 1000 wild-type sequences using PCR products without the need of LDR amplification. Additionally, this on-chip beads ligation approach also displayed other microfluidic-based advantages of simple handling (one sample injection per test), little reagent quantities, and low potential of contaminations.<br />
<br />
<br />
<br />
<br />
<br />
'''Link:''' [http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6TFC-4PSC23F-3&_user=1067472&_rdoc=1&_fmt=&_orig=search&_sort=d&_docanchor=&view=c&_searchStrId=1058201776&_rerunOrigin=scholar.google&_acct=C000051251&_version=1&_urlVersion=0&_userid=1067472&md5=7df5e0a61ca6e430d450c78873bafd66 Science Direct]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Microfabricated_Device_for_DNA_and_RNA_Amplification_by_Continuous-Flow_Polymerase_Chain_Reaction_and_Reverse_Transcription-Polymerase_Chain_Reaction_with_Cycle_Number_SelectionTeam:Alberta/References/Publications/Microfabricated Device for DNA and RNA Amplification by Continuous-Flow Polymerase Chain Reaction and Reverse Transcription-Polymerase Chain Reaction with Cycle Number Selection2009-10-21T17:22:42Z<p>Mitch p: </p>
<hr />
<div>Authors: Pierre J. Obeid, Theodore K. Christopoulos, H. John Crabtree, and Christopher J. Backhouse<br />
<br />
Anal. Chem., 2003, 75 (2), pp 288–295<br />
<br />
<br />
<b>Abstract:</b> We have developed a high-throughput microfabricated, reusable glass chip for the functional integration of reverse transcription (RT) and polymerase chain reaction (PCR) in a continuous-flow mode. The chip allows for selection of the number of amplification cycles. A single microchannel network was etched that defines four distinct zones, one for RT and three for PCR (denaturation, annealing, extension). The zone temperatures were controlled by placing the chip over four heating blocks. Samples and reagents for RT and PCR were pumped continuously through appropriate access holes. Outlet channels were etched after cycles 20, 25, 30, 35, and 40 for product collection. The surface-to-volume ratio for the PCR channel is 57 mm-1 and the channel depth is 55 μm, both of which allow very rapid heat transfer. As a result, we were able to collect PCR product after 30 amplification cycles in only 6 min. Products were collected in 0.2-mL tubes and analyzed by agarose gel electrophoresis and ethidium bromide staining. We studied DNA and RNA amplification as a function of cycle number. The effect of the number of the initial DNA and RNA input molecules was studied in the range of 2.5 × 106−1.6 × 108 and 6.2 × 106−2 × 108, respectively. Successful amplification of a single-copy gene (β-globin) from human genomic DNA was carried out. Furthermore, PCR was performed on three samples of DNA of different lengths (each of 2-μL reaction volume) flowing simultaneously in the chip, and the products were collected after various numbers of cycles. Reverse transcription was also carried out on four RNA samples (0.7-μL reaction volume) flowing simultaneously in the chip, followed by PCR amplification. Finally, we have demonstrated the concept of manually pumped injection and transport of the reaction mixture in continuous-flow PCR for the rapid generation of amplification products with minimal instrumentation. To our knowledge, this is the first report of a monolithic microdevice that integrates continuous-flow RT and PCR with cycle number selection. <br />
<br />
<br />
<br />
<br />
'''Link:''' [http://pubs.acs.org/doi/abs/10.1021/ac0260239 ACS Publications]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Microelectromagnet_for_magnetic_manipulation_in_lab-on-a-chip_systemsTeam:Alberta/References/Publications/Microelectromagnet for magnetic manipulation in lab-on-a-chip systems2009-10-21T17:18:16Z<p>Mitch p: </p>
<hr />
<div>Authors:Kristian Smistrupa, Peter T. Tangb, Ole Hansena and Mikkel F. Hansen <br />
<br />
Journal of Magnetism and Magnetic Materials Volume 300, Issue 2, May 2006, Pages 418-426 <br />
<br />
<br />
<b>Abstract:</b> We demonstrate a simple scheme for fabrication of microelectromagnets consisting of planar spiral coils semi-encapsulated in soft magnetic yokes using conventional microfabrication techniques. The microelectromagnets are suitable for applications operating at frequencies below 250 kHz. Conventional fabrication schemes for planar microelectromagnets typically rely on five mask steps. We allow the current to flow in the soft magnetic yoke and thereby two mask steps are eliminated. We have characterized the electromagnets electrically, the results agree well with theory, and the implications arising from current flowing in the magnetic yoke are discussed. We have integrated the microelectromagnets with microfluidic channels, and demonstrated separation of commercially available magnetic beads from a fluid in a microfluidic system, i.e. a lab-on-a-chip system.<br />
<br />
<br />
<br />
<br />
<br />
'''Link:''' [http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6TJJ-4GFN3YH-6&_user=1067472&_rdoc=1&_fmt=&_orig=search&_sort=d&_docanchor=&view=c&_searchStrId=1058191490&_rerunOrigin=google&_acct=C000051251&_version=1&_urlVersion=0&_userid=1067472&md5=a8edb9d14d3de834edac7c070f0b3cbf Science Direct]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/An_integrated_microfluidic_biochemical_detection_system_forprotein_analysis_with_magnetic_bead-based_samplingcapabilitiesTeam:Alberta/References/Publications/An integrated microfluidic biochemical detection system forprotein analysis with magnetic bead-based samplingcapabilities2009-10-21T17:12:38Z<p>Mitch p: </p>
<hr />
<div>Authors: Choi JW, Oh KW, Thomas JH, Heineman WR, Halsall HB, Nevin JH, Helmicki AJ, Henderson HT, Ahn CH. <br />
<br />
Lab Chip 2002. Feb; 2(1): 27-30<br />
<br />
<br />
<b>Abstract:</b> This paper presents the development and characterization of an integrated microfluidic biochemical detection system for fast and low-volume immunoassays using magnetic beads, which are used as both immobilization surfaces and bio-molecule carriers. Microfluidic components have been developed and integrated to construct a microfluidic biochemical detection system. Magnetic bead-based immunoassay, as a typical example of biochemical detection and analysis, has been successfully performed on the integrated microfluidic biochemical analysis system that includes a surface-mounted biofilter and electrochemical sensor on a glass microfluidic motherboard. Total time required for an immunoassay was less than 20 min including sample incubation time, and sample volume wasted was less than 50 microl during five repeated assays. Fast and low-volume biochemical analysis has been successfully achieved with the developed biofilter and immunosensor, which is integrated to the microfluidic system. Such a magnetic bead-based biochemical detection system, described in this paper, can be applied to protein analysis systems.<br />
<br />
<br />
<br />
<br />
<br />
'''Link:''' [http://www.ncbi.nlm.nih.gov/pubmed/15100857 PubMed]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Magnetic_bead_handling_on-chip:_new_opportunities_for_analytical_applicationsTeam:Alberta/References/Publications/Magnetic bead handling on-chip: new opportunities for analytical applications2009-10-21T17:11:49Z<p>Mitch p: </p>
<hr />
<div>Authors: Gijs, Martin<br />
<br />
Microfluidics and Nanofluidics, Volume 1, Number 1, November 2004 , pp. 22-40(19)<br />
<br />
<br />
<b>Abstract:</b> This review describes recent advances in the handling and manipulation of magnetic particles in microfluidic systems. Starting from the properties of magnetic nanoparticles and microparticles, their use in magnetic separation, immuno-assays, magnetic resonance imaging, drug delivery, and hyperthermia is discussed. We then focus on new developments in magnetic manipulation, separation, transport, and detection of magnetic microparticles and nanoparticles in microfluidic systems, pointing out the advantages and prospects of these concepts for future analysis applications. <br />
<br />
<br />
<br />
'''Link:''' [http://www.ingentaconnect.com/content/klu/10404/2004/00000001/00000001/art00003 IngentaConnect]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/Development_of_a_microfluidic_biosensor_module_for_pathogen_detectionTeam:Alberta/References/Publications/Development of a microfluidic biosensor module for pathogen detection2009-10-21T17:08:34Z<p>Mitch p: </p>
<hr />
<div>Authors: Natalya V. Zaytsevaa, Vasiliy N. Goralb, Richard A. Montagnab and Antje J. Baeumner<br />
<br />
Lab Chip, 2005, 5, 805-811<br />
<br />
<br />
<br />
<b>Abstract:</b> The development of a microfluidic biosensor module with fluorescence detection for the identification of pathogenic organisms and viruses is presented in this article. The microfluidic biosensor consists of a network of microchannels fabricated in polydimethylsiloxane (PDMS) substrate. The microchannels are sealed with a glass substrate and packed in a Plexiglas housing to provide connection to the macro-world and ensure leakage-free flow operation. Reversible sealing permits easy disassembly for cleaning and replacing the microfluidic channels. The fluidic flow is generated by an applied positive pressure gradient, and the module can be operated under continuous solution flow of up to 80 µL min–1. The biosensor recognition principle is based on DNA/RNA hybridization and liposome signal amplification. Superparamagnetic beads are incorporated into the system as a mobile solid support and are an essential part of the analysis scheme. In this study, the design, fabrication and the optimization of concentrations and amounts of the different biosensor components are carried out. The total time required for an assay is only 15 min including sample incubation time. The biosensor module is designed so that it can be easily integrated with a micro total analysis system, which will combine sample preparation and detection steps onto a single chip.<br />
<br />
<br />
<br />
<br />
<br />
'''Link:''' [http://www.rsc.org/delivery/_ArticleLinking/DisplayHTMLArticleforfree.cfm?JournalCode=LC&Year=2005&ManuscriptID=b503856a&Iss=8 RSC Publishing]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/An_integrated_microfluidic_biochemical_detection_system_forprotein_analysis_with_magnetic_bead-based_samplingcapabilitiesTeam:Alberta/References/Publications/An integrated microfluidic biochemical detection system forprotein analysis with magnetic bead-based samplingcapabilities2009-10-21T17:04:29Z<p>Mitch p: </p>
<hr />
<div>Authors: Choi JW, Oh KW, Thomas JH, Heineman WR, Halsall HB, Nevin JH, Helmicki AJ, Henderson HT, Ahn CH. <br />
<br />
Lab Chip 2002. Feb; 2(1): 27-30<br />
<br />
<br />
<b>Abstract:</b> This paper presents the development and characterization of an integrated microfluidic biochemical detection system for fast and low-volume immunoassays using magnetic beads, which are used as both immobilization surfaces and bio-molecule carriers. Microfluidic components have been developed and integrated to construct a microfluidic biochemical detection system. Magnetic bead-based immunoassay, as a typical example of biochemical detection and analysis, has been successfully performed on the integrated microfluidic biochemical analysis system that includes a surface-mounted biofilter and electrochemical sensor on a glass microfluidic motherboard. Total time required for an immunoassay was less than 20 min including sample incubation time, and sample volume wasted was less than 50 microl during five repeated assays. Fast and low-volume biochemical analysis has been successfully achieved with the developed biofilter and immunosensor, which is integrated to the microfluidic system. Such a magnetic bead-based biochemical detection system, described in this paper, can be applied to protein analysis systems.<br />
<br />
<br />
<br />
<br />
<br />
'''Link:''' [http://http://www.ncbi.nlm.nih.gov/pubmed/15100857 PubMed]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/An_inexpensive_and_portable_microchip-based_platform_for_integratedRT%E2%80%93PCR_and_capillary_electrophoresisTeam:Alberta/References/Publications/An inexpensive and portable microchip-based platform for integratedRT–PCR and capillary electrophoresis2009-10-21T17:00:01Z<p>Mitch p: </p>
<hr />
<div>Authors: <br />
Govind V Kaigala, Viet N Hoang, Alex Stickel, Jana Lauzon, Dammika Manage, Linda M Pilarski, Christopher J Backhouse<br />
<br />
The Analyst 133(3):331-8 01/04/2008 <br />
<br />
<br />
<b>Abstract:</b> We present an inexpensive, portable and integrated microfluidic instrument that is optimized to perform genetic amplification and analysis on a single sample. Biochemical reactions and analytical separations for genetic analysis are performed within tri-layered glass-PDMS microchips. The microchip itself consists of integrated pneumatically-actuated valves and pumps for fluid handling, a thin-film resistive element that acts simultaneously as a heater and a temperature sensor, and channels for capillary electrophoresis (CE). The platform is comprised of high voltage circuitry, an optical assembly consisting of a laser diode and a charged coupled device (CCD) camera, circuitry for thermal control, and mini-pumps to generate vacuum/pressure to operate the on-chip diaphragm-based pumps and valves. Using this microchip and instrument, we demonstrate an integration of reverse transcription (RT), polymerase chain reaction (PCR), and capillary electrophoresis (CE). The novelty of this system lies in the cost-effective integration of microfluidics, optics, and electronics to realize a fully portable and inexpensive system (on the order of $1000 in component costs) for performing both genetic amplification and analysis - the basis of many medical diagnostics. We believe that this combination of portability, cost-effectiveness and performance will enable more accessible healthcare.<br />
<br />
<br />
<br />
<br />
'''Link:''' [https://www.researchgate.net/publication/5552096_An_inexpensive_and_portable_microchip-based_platform_for_integrated_RT-PCR_and_capillary_electrophoresis ResearchGATE]</div>Mitch phttp://2009.igem.org/Team:Alberta/References/Publications/An_inexpensive_and_portable_microchip-based_platform_for_integratedRT%E2%80%93PCR_and_capillary_electrophoresisTeam:Alberta/References/Publications/An inexpensive and portable microchip-based platform for integratedRT–PCR and capillary electrophoresis2009-10-21T16:57:57Z<p>Mitch p: </p>
<hr />
<div>Authors: <br />
Govind V Kaigala, Viet N Hoang, Alex Stickel, Jana Lauzon, Dammika Manage, Linda M Pilarski, Christopher J Backhouse<br />
<br />
The Analyst 133(3):331-8 01/04/2008 <br />
<br />
<br />
<b>Abstract:</b> We present an inexpensive, portable and integrated microfluidic instrument that is optimized to perform genetic amplification and analysis on a single sample. Biochemical reactions and analytical separations for genetic analysis are performed within tri-layered glass-PDMS microchips. The microchip itself consists of integrated pneumatically-actuated valves and pumps for fluid handling, a thin-film resistive element that acts simultaneously as a heater and a temperature sensor, and channels for capillary electrophoresis (CE). The platform is comprised of high voltage circuitry, an optical assembly consisting of a laser diode and a charged coupled device (CCD) camera, circuitry for thermal control, and mini-pumps to generate vacuum/pressure to operate the on-chip diaphragm-based pumps and valves. Using this microchip and instrument, we demonstrate an integration of reverse transcription (RT), polymerase chain reaction (PCR), and capillary electrophoresis (CE). The novelty of this system lies in the cost-effective integration of microfluidics, optics, and electronics to realize a fully portable and inexpensive system (on the order of $1000 in component costs) for performing both genetic amplification and analysis - the basis of many medical diagnostics. We believe that this combination of portability, cost-effectiveness and performance will enable more accessible healthcare.<br />
<br />
<br />
<br />
<br />
'''Link:''' [http://eeeeeeeeeeeee NAME]</div>Mitch phttp://2009.igem.org/Team:AlbertaTeam:Alberta2009-10-21T13:37:53Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/Template3}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#e1e1e1; margin:0 5px;}<br />
.b2f {height:1px; background:#e1e1e1; margin:0 3px;}<br />
.b3f {height:1px; background:#e1e1e1; margin:0 2px;}<br />
.b4f {height:2px; background:#e1e1e1; margin:0 1px;}<br />
.content {background: #e1e1e1;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=60% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 200; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Recoli"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>BioBytes</h1><br />
<br />
<div align="justify"><br />
<br />
<font size="2"><br />
<p>At present, the cost to synthesize oligonucleotides has been continually declining and therefore their availability is exponentially growing. However, the current technique to ligate pieces of DNA together is outdated. Present methods take a considerable amount of time to piece DNA together, making large constructs incredibly difficult to build. The University of Alberta 2009 iGEM team would like to introduce the BioBytes Chromosome Assembly System. This method refers to a mechanism for rapid and reliable construction of plasmids (i.e. artificial gene sets) in vitro. It allows for the assembly of components in a structured manner within hours rather than days. Our hope is to see this method become a valuable tool for any molecular biologist. Furthermore we have adapted our approach for Biofabrication by developing a robot which can use our method for automated assembly. Finally, microfluidics have been utilized to miniaturize construction allowing for additional validation for automated production of constructs.</p><br />
<p align=right><p align=right><a href="https://2009.igem.org/Team:Alberta/Project/assemblyoverview"> Click here for more...</a> </P><br />
<p>The method can be applied to numerous different applications, however, its greatest application is for the assembling of entire genomes. For this reason we have provided a detailed explanation regarding the requirements of constructing a minimal genome including an in-silico method for identifying essential genes in any organism, and a theoretical design of replacing the host chromosome with the new synthetic genome.</p><br />
<p align=right><p align=right><a href="https://2009.igem.org/Team:Alberta/Project/Bioinformatics"> Click here for more...</a> </P><br />
<br />
</font></div><br />
<br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td style="height: 800; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Overview"><br />
<div style="height: 800; background:#FFFFFF; line-height:100% padding: 3px 0px;"><br />
<h2>The Minimal Genome Project</h2><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<font size="2"><P>The minimal <i>Escherichia coli</i> genome has been the holy grail of biology for a number of years. <i>E. coli</i> is the most widely used cellular research tool by the molecular biology community. Since scientific research is based upon reductionism and simplification for understanding, a simplified version of an experimental model organism such as <i>E. coli</i> is ideally preferred as a chassis for experimentation. Reducing the <i>E. coli</i> genome to roughly 10% of its original size, demonstrates a great simplification of this model organism.</P><br />
<P><br />
To reconstruct such an organism, we plan on building an artificial <i>E. coli</i> chromosome using the BioBytes chromosome assembly system and inserting it into living <i>E. coli</i> cells. We then intend to remove the host chromosome by making it incapable of division, thus allowing only the artificially inserted chromosome to propagate through multiple generations as the cells grow and divide. This is markedly different from the current, time-consuming method of knocking out inessential genes, one at a time, in an effort to produce the minimal genome. It is this difference that we hope to exploit in our attempt to win the race to produce the minimal <i>E. coli</i> genome.</p><br />
</font><br />
</div><br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
<br />
</td><br />
</tr><br />
<tr><br />
<td style="height: 800; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Overview"><br />
<div style="height: 800; background:#FFFFFF; line-height:100% padding: 3px 0px;"><br />
<h2>Team Achievements</h2><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<font size="2"><P> Through our efforts we have made the following accomplishments:</p><br />
<ul><br />
<li>Developed the BioBytes Assembly Method and produced a proof of concept of the design<br />
<li>A kit of components has been added to the registry allowing for use of our system by anyone<br />
<li>Produced a series of modeling programs which can be used to determine the essential genes in any organism<br />
<li>188 essential genes have been amplified and their primers added to the registry<br />
<li>A robot has been developed demonstrating the potential of automation for BioBytes<br />
<li>Used microfluidics to show the biofabrication potential of our design<br />
<li>We have hosted a debate involving synthetic biology<br />
<li>We have constructed and completed a series of presentation to discuss iGEM and promote knowledge of synthetic biology <br />
</ul><br />
</font><br />
</div><br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
<br />
</td><br />
</tr><br />
<br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/Team:Alberta/TeamTeam:Alberta/Team2009-10-20T21:50:55Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/Template3}}<br />
<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#e1e1e1; margin:0 5px;}<br />
.b2f {height:1px; background:#e1e1e1; margin:0 3px;}<br />
.b3f {height:1px; background:#e1e1e1; margin:0 2px;}<br />
.b4f {height:2px; background:#e1e1e1; margin:0 1px;}<br />
.content {background: #e1e1e1;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=60% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Recoli"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>Project BioBytes</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/6/67/UofA_iGEM2009_IMG_5009.JPG" width="320" height="407"><br />
</center><br />
<br />
<br> <br />
<br />
<table><br />
<br />
<tr><br />
<td><br />
<div align="left"><strong>Kalon Armstrong</strong> <br><br />
<em>Molecular Genetics</em> <br><br />
</div><br />
<td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/3/34/UofA09_team_Kalon_Armstrong.jpg" hspace="20"><br />
</td><br />
<td><br />
<div align="justify" >I've recently completed my BSc in Molecular Genetics and will be entering the Engineering program in the fall of 2009. Most of my growing-up took place in the small town of Cochrane, Alberta. My ultimate goals consist of working in the Biotechnology or health care industry. While my interest in music, movies, snowboarding, and hanging out with friends may seem stereotypical on the surface, they feel unique in their own right and keep me busy most of the time. This will be my first year on the U of A iGEM team and I am excited to help take the competition to a new level. I think that iGEM will be a great experience because it demands innovation and collaboration on levels rarely seen in undergraduate programs.</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td colspan = "2"><br />
<br />
<hr style=width:66% align=left><br />
</td><br />
<tr><br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Eric Bennett</strong><br><br />
<em>Electrical Biomedical Engineering</em><br><br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/7/7a/UofA09_team_Eric_Bennett.jpg" hspace="20"><br />
</td><br />
<br />
<td><br />
<div align="justify" >I am entering my final year of engineering at the U of A. After graduation, I hope to do research either with a biotechnology company or in graduate level studies. I think that iGEM is a great way to gain valuable experience and is an effective way of accelerating the field of synthetic biology. My interests include brain-machine interfacing, genetic engineering, and robotic control systems. My hobbies include playing guitar, video games, the occasional sport, reading, and fixing my car.</div><br />
</td><br />
</tr> <br />
<br />
<tr><br />
<td colspan = "2"><br />
<br />
<hr style=width:66% align=left><br />
</tr><br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Max Buchko</strong><br><br />
<em>Honors Biochemistry</em><br><br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/e/e1/UofA09_team_Max_Buchko.jpg" ALIGN="LEFT" hspace="20"><br />
</td><br />
<br />
<td><br />
<div align="justify" style="padding-right:20px">I am in my third year of Honors Biochemistry and wish to pursue a career in medicine. In my time away from the lab I enjoy a wide variety of sports including soccer, boxing, and rifle silhouette shooting. I have also been known to strum a chord or two at an intolerable volume to the annoyance of the people living upstairs.<br />
I hope for the best with iGEM at MIT in 2009. This competition is a means of proving yourself at an exemplary level amongst many of the top international minds, and it is this challenge that I look forward to the most.</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td colspan = "2"><br />
<br />
<hr style=width:66% align=left><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Oscar Cortes</strong><br><br />
<em>Bsc. Specialization molecular genetics</em><br><br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/7/70/UofA09_team_Oscar_Cortes.jpg" ALIGN="LEFT" hspace="20"><br />
</td><br />
<td><br />
<div align="justify" style="padding-right:20px">My future plans include entering into a Masters program in Medical Genetics or Human Genetics, and pursuing this discipline towards a PhD. I enjoy reading books about the human genome and advancements in stem cell research. In my spare time I like to play a variety of sports that I am not necessarily good at: soccer, softball, and dodge ball. I have also been known to pound on a drum set from time to time without any rhythm whatsoever. I see iGEM as an extraordinary opportunity for me to be exposed to real life research, in which my knowledge of molecular genetics will be challenged, and will help me further my understanding of Synthetic Biology.<br />
"Man with all his noble qualities still bears in his bodily frame the indelible stamp of his lowly origin"- Charles Darwin</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td colspan = "2"><br />
<br />
<hr style=width:66% align=left><br />
</td><br />
</tr> <br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Anh Dao</strong><br><br />
<em>Biological Sciences and Chemistry</em><br> <br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/c/cf/UofA09_team_Ahn_Dao.jpg" ALIGN="LEFT" hspace="20"><br />
</td><br />
<br />
<td><br />
<div align="justify" style="padding-right:20px">I am interested in a research career after I finish my degree. I am leaning towards the field of Microbiology to study the many micro-organisms that have not yet been discovered. However, I may enroll in graduate studies after my undergraduate degree to expand my knowledge and gain more experience in the laboratory. Being in a competitive team and atmosphere is a motivating and exciting opportunity that I do not want to miss out on. By creating the smallest artificial E. coli genome we can extend future research. We are attempting to understand and standardize the E. coli genome so that this methodology can be applied to more complex model organisms.</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Uchechukwu Davidson</strong><br><br />
<em>Honors biochemistry</em><br><br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/3/3c/UofA09_team_Uche_Davidson.jpg" ALIGN="LEFT" hspace="20"><br />
</td><br />
<td><br />
<div align="justify" style="padding-right:20px">I am in my third year of Biochemistry at the University of Alberta. I wish to pursue a career in medicine after my degree. I enjoy sports, movies, and music at my time away from my studies. The iGEM provides an opportunity to experience creativity, innovation, and ingenuity which are sometimes absent at the undergraduate level of science.</div><br />
</td><br />
</tr><br />
<br />
<tr> <br />
<td colspan = "2"><br />
<br />
<hr style=width:66% align=left><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Youness Elkhalidy</strong><br><br />
<em>Honors immunology and infection</em><br><br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/f/fe/UofAiGEM2009_Youness_IMG_5049.JPG" width="200" height="300" ALIGN="LEFT" hspace="20"><br />
</td><br />
<td><br />
I am a first year student at the University of Alberta. I hope to enter medical school in the near future. I am currently taking part in mitotic-spindle regulation research from a genetics perspective. I have a passion for science and enjoy sports such as basketball and soccer. iGEM is an great learning experience not only in the cutting-edge field of Synthetic Biology but also in leadership and business management. I will enjoy taking part in research that combines many fields of science as well as socialize with many of my team members who share common interests.<br />
</td><br />
</tr><br />
<br />
<tr><br />
<td colspan = "2"><br />
<br />
<hr style=width:66% align=left><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Justin Fedor</strong><br><br />
<em>Honours Biochemistry</em><br><br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/7/71/UofA09_team_Justin_Fedor.jpg" ALIGN="LEFT" hspace="20"><br />
</td><br />
<br />
<td><br />
<div align="justify" style="padding-right:20px">I have completed my undergraduate degree in Biochemistry this year and will be starting my Ph.D. program in September. I play piano and am attempting to learn the cello. My nerdy tendencies are obvious when I say that I like Star Trek TNG but not DS9. Last summer I worked in the biomedical research lab of Dr. Larry Unsworth of the National Institute for Nanotechnology (NINT), which has further piqued my interest in the field of nanotechnology. In the hopefully not too distant future I plan on becoming a researcher studying the mechanisms of membrane bound enzymes, particularly oxidoreductases.</div><br />
</td><br />
</tr><br />
<br />
<tr><td colspan = "2"> <br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Jason Gardiner</strong> <br><br />
<em>BSc. Specialization in Botany</em><br><br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/2/2b/UofA09_team_Jason_Gardiner.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">Jason is a veteran of iGEM 2007, where his team "The Butanerds" won first place in the Energy track. Jason is currently in his fourth year of his Bachelor degree specializing in Botany. His hobbies include Softball, Beach volleyball, soccer, as well as playing the guitar and fiddling with the iGEM 2009 Wiki. Jason has applied to continue his education in graduate studies at the University of Alberta in 2010. His degree in Botany focuses mostly on the molecular side of plants and he hopes to use this knowledge applying Synthetic Biology to plants. One day he hopes to solve all of the worlds problems using plants and Synthetic Biology.</div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><br />
<div align="left"><br />
<strong>Erin Garside</strong> <br><br />
<em>Biological Sciences</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/6/60/UofA09_team_Erin_Garside.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">I completed my BSc. degree this year and plan to do graduate work in biochemistry. After that - who knows? I think iGEM will be a great experience, and that it's about time Synthetic Biology really took off. In my spare time I like to read, play computer games (especially the Sims) and raise cats.</div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Boris Henriquez</strong><br><br />
<em>Biological Sciences</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/e/ec/UofA09_team_Boris_Heneriquez.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I have just completed the third year of my BSc. degree. My favorite classes so far have been in the fields of Biochemistry and Genetics. I am most interested in human development and embryogenesis and hope to learn more about these topics in my fourth year. My ultimate goal is to have a career in the medical industry as both a physician and as a researcher. When I'm not busy with school or work my usual activities are pretty standard. I like to hang out with friends and family and I crave the outdoors. This is my first year with iGEM and I'm really excited to learn more about Synthetic Biology. </div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Stephen Jahns</strong><br><br />
<em>Molecular Genetics</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/a/a8/Alberta_steve.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I am in my second year at the University of Alberta. After taking a year of engineering, I decided to transfer to molecular genetics in order to fulfill my childhood desire of creating an army of giant bat-rabbits. I plan on going on through to graduate school to earn a PhD in order to reach my goals. Also, health research could be pretty interesting too. In my spare time I like to run around, ride my bike, play music, cook delicious food, and pretty much all the other fun things kids are doing these days. In my first year I helped to build a car with the U of A's Formula SAE fabrication team. I had a good time building stuff out of metal, and this year I know that I'm going to have a blast building (or at least attempting to build) a novel organism out of chunks of DNA. I have a lot to learn but I know it will be worth it. </div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<br />
<tr><td><div align="left"><br />
<strong>Eric Leung</strong> <br><br />
<em>Honors Pharmacology</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/a/a1/UofA09_team_Eric_Leung.jpg" ALIGN="LEFT" hspace="20"><br />
<br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">I am currently in my fourth and final year of an Honors Pharmacology degree. What I do after this degree is up in the air but it is definitely something in the health sciences. I hope I can become a professional baker in my spare time. Traveling to Europe, Mexico, and Japan is also in the books somewhere along the way.</div><br />
<br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<br />
<tr><td><div align="left"><br />
<strong>David Lloyd</strong><br> <br />
<em> Biochemistry</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/6/6f/UofA09_team_David_Lloyd.jpg" ALIGN="LEFT" hspace="20"><br />
<br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">I am a fourth year student, studying at the University of Alberta. While Biochemistry is my major, I have many interests including Genetics, Immunology, Cell Biology, Bioinformatics, and Microbiology. In my spare time I can be found playing sports like soccer, volleyball, or squash, as well as playing piano, listening to music, and playing video games. In the future, I hope to continue into a graduate program in Biochemistry, Cell Biology, Synthetic Biology, or in another field. iGEM is of great interest to myself because of its application to the future of Synthetic Biology. It is an awesome challenge which will hopefully help me to cultivate myself.</div><br />
<br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Enoch Ng</strong><br><br />
<em>Biological Sciences/Business</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/7/7c/UofA09_team_Enoch_Ng.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I am entering my fourth year of studies at the University of Alberta, and am looking forward to the challenge that iGEM provides. In the future I hope to travel across the world and meet people ranging from Meshaal to Obama and remain a glorified generalist. </div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Emera Nguyen</strong><br><br />
<em>BSc. Biological Sciences/Economics</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/5/52/UofA09_team_Emera_Nguyen.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I am a BSc student entering my fourth year in a Biological Sciences major and Economics minor. This year will be my second year on an iGEM team, and my first year representing the U of A team. In my downtime, things I enjoy include spending quality time with friends and family and traveling to lesser-known parts of the world. One year's experience later, the iGEM competition still impresses me with the unique opportunities it provides for student growth. This research competition is one of the highlights of my undergraduate experience.</div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr> <br />
<br />
<br />
<tr><td><div align="left"><br />
<strong>Mitch Paquette</strong><br><br />
<em>BSc. Honors Molecular Genetics</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/b/bf/Mitch5012_uofa.igem09.JPG" ALIGN="LEFT" hspace="20"><br />
<br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">I have transfered into the honors program in Molecular Genetics from a BSc General program in physical science. After my degree is completed I plan to apply to graduate studies in Molecular Biology, after which I hope to work in research or academia. What excites me most about iGEM is the opportunity to gain research experience in such a new field.</div><br />
</td></tr> <br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Amber Paul</strong><br><br />
<em>BSc. Specialization Immunology and Infection</em><br><br />
</div></td></tr><br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/3/3b/UofA_Team_Amber.jpg" width="200" height="300" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I plan to find the cure for cancer. Seriously. Will be attending graduate school to earn a PhD following my undergraduate degree. I would like to live somewhere warm that allows me to do research, preferably Maui. iGEM is a developmental forefront to Synthetic Biology, something that I love to be part of. Its challenging and fun. To be able to determine the minimal genes for a simple prokaryote, we provide a scaffold to future research on larger, more complex cells such as cancerous tissues, simple eukaryotes, etc.</div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Julia Pon</strong><br><br />
<em>Honors Molecular Genetics</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/3/30/UofA09_team_Julia_Pon.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">My future plans include pursuing PhD in Medical Genetics. I've worked in biological research labs for the past two summers, with last summer spent at the German Cancer Research Institute in Heidelberg, Germany. I'll be working in both the iGEM lab and a Medical Genetics research lab during summer 2009. My heritage is a mixture of English, Chinese, and Danish. iGEM helps move biology to a streamlined, standardized, abstracted process that opens the door to interdisciplinary collaborations and new applications. I'm excited about the many applications of Synthetic Biology and am enjoying the student-directed and team-oriented nature of iGEM.</div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<br />
<tr><td><div align="left"> <br />
<strong>Alina Ponomarev</strong><br><br />
<em>Biological Sciences</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/e/e1/UofA2009iGEM_Alina_IMG_5023.JPG" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I am currently a second year student in Biological Sciences and I am considering transferring into the Immunology and Infection program for my third year. I joined iGEM at the end of my first year and can honestly say that it has been an unbeatable learning experience. It is very exciting to think that the progress we have made can positively impact synthetic biology and society in general. In the future, my career goal is to become a Pediatrician. However, after doing labwork at the iGEM lab this summer, I have begun to consider a career in medical research. </div> <br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Kelly Robinson</strong><br><br />
<em>Hon. Biochemistry</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/9/92/UofA09_team_Kelly_Robinson.png" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">I completed my BSc. in Biochemistry this year and have enrolled in the University of Alberta's Chemical Engineering program in hopes of fusing the fundamental science of biological machines with the entrepreneurial world of engineering. I got involved in iGEM a couple of years ago when I went to a presentation put on by one of the past teams. I ended up applying in the middle of the night on the last day of registration. That said, it has been a great experience ever since. I'm glad that this year I get the opportunity to do some full time work and really get into the project. Overall, iGEM has a lot to offer anyone who gets involved (including incorrigible advisors: you know who you are!).</div><br />
<br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr> <br />
<br />
<tr><td><div align="left"><br />
<strong>James Rodway</strong><br><br />
<em>Electrical Engineering</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/a/a5/UofA09_team_James_Rodway.jpg" ALIGN="LEFT" hspace="20"><br />
<br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">I am currently finishing up an Electrical Engineering degree, and am aiming to do some graduate work in more of the computer area, specifically modeling. I've worked at a few cool places during co-op work terms and have done things like environmental monitoring and a whole mess of smaller projects during my time in the 'cube farm'. At the University I've been involved with the ARVP and iGEM, as best as time has allowed me.<br />
I haven't really had too much time for hobbies in the last while, but when I did I played more video games, the guitar, and I actually read books for entertainment value. During my time on last year's iGEM team I found that it was a pretty cool multidisciplinary project, which drew in a few different disciplines that I have never really interacted with at all previously. It was very impressive to see what we, and all the iGEM teams, had accomplished by the end of last year's competition.</div><br />
<br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Andy Spencer</strong><br><br />
<em>Honours Biochemistry</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/c/ca/UofA09_team_Andy_Spencer.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">In 5 years I see myself on one of two different paths. I will start applying to medical colleges this summer. Alternatively, I am interested in discovering novel treatments for cancer, and am considering a career in Research Oncology. I grew up in the Okanagan, British Columbia, and love spending my summers at the beach . In my spare time I enjoy playing squash and bouldering. iGEM to me represents a fundamental value of science as a discipline; that people can from diverse backgrounds with a common interest and curiosity can work as a cohesive team to overcome barriers and problems. I am excited to meet iGEM participants from around the world, and to visit MIT in November.</div><br />
<br />
</td></tr> <br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Jonathan Tam</strong><br><br />
<em>Honors Cell Biotechnology</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/e/e2/UofA09_team_Jonathan_Tam.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I have completed my Bachelor's degree in Honors Cell Biotechnology and intend to pursue a medical degree in Germany in the near future. I am very interested in the fields of Molecular Biology and Immunology. For the last two years, I have been keeping myself busy studying the development of macrophages in the lab of Dr. Daniel Barreda. Outside of the lab, I am an avid photographer and downhill mountain biker. The iGEM competition accelerates the development of Synthetic Biology as a field. To be a part of our University of Alberta team is an excellent opportunity for collaboration and advancement.</div><br />
</td></tr> <br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Jennifer Yau</strong><br><br />
<em>Honors Biochemistry</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/a/ac/UofA09_team_Jennifer_Yau.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I am currently in the second year of my undergraduate program with the ultimate goal of attending graduate school to pursue a research career in Geriatric Medicine. Additional science related activities of mine include my interest in astronomy and promoting science to elementary students through volunteer work. Otherwise, I dedicate the majority of my free time to the arts of chainmaille, knitting, and jazz/classical piano. This is my first year on iGEM and I am ecstatic to be partaking in a project that requires such diverse disciplinary backgrounds. Not only will this be a great learning experience in so many perspectives, but also an opportunity to contribute significant ideas to the ongoing research in Synthetic Biology</div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr> <br />
<br />
<tr><td><div align="left"><br />
<strong>Zach Wiltshire</strong> <br><br />
<em>BSc. Specialization in Cell Biology</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/1/19/UofA09_team_Zach_Wiltshire.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">I am entering into my fourth year in the Cell Biology program at the U of A and have already experienced iGEM once before as a member of the 2008 National Institute for Nanotechnology team. Through my degree I have found myself to have interests ranging from immunology to both eukaryotic & prokaryotic cellular anatomy & physiology. I also happen to be very fond of molecules that fluoresce. My experience with iGEM last year opened my eyes to the possibilities which engineering biological systems can offer. Over the course of the 2009 competition I hope to have just as many opportunities to learn as I did last year.</div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<tr><td><br />
<a name="Team_Ad"></a><br />
<strong>The Supervisors</strong><br />
</tr><td><br />
<br />
<tr><td><br />
<strong>Mike Ellison</strong><br />
<img src="https://static.igem.org/mediawiki/2009/d/d5/UofA09_teamad_Mike_Ellison.jpg" ALIGN="LEFT" hspace="20"><br />
</td></tr><br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><br />
<br />
<strong>Doug Ridgway</strong><br />
<img src="https://static.igem.org/mediawiki/2009/c/cc/UofA09_teamad_Douglas_Ridgway.jpg" ALIGN="LEFT" hspace="20"></a><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><br />
<strong>James Maclagan</strong><br />
<img src="https://static.igem.org/mediawiki/2009/a/ab/UofAigem2009_JamesMIMG_4975.JPG" ALIGN="LEFT" hspace="20"></a><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</tr></td><br />
<tr><td><br />
<a name="Fac_Con"></a><br />
<strong>Faculty Consultants</strong><br />
</tr><td><br />
<br />
<tr><td><br />
<strong>Chris Backhouse</strong><br />
<br><br />
Department of Electrical Engineering<br />
<br><br />
christopher.backhouse@ualberta.ca<br />
<img src="http://www.ece.ualberta.ca/~chrisb/Graphics/Backhouse.jpg" ALIGN="LEFT" hspace="20"><br />
</td><br />
<br />
<td><br />
<div align="justify" ><br />
Nanobiotechnologies give us the ability to manipulate and sense at the level of individual molecules, with a tremendous potential impact on both human health and the economy. To a large extent, this potential is likely to be realised through the development of Lab on Chip (LOC) technologies. Although the LOC technologies are powerful, the complexity of the infrastructure required to support LOC operation has hindered the widespread adoption of LOC methods in life science applications. A central theme in the work of the Backhouse lab (<a href="http://www.ece.ualberta.ca/~aml/">the Applied Miniaturisation Lab, AML</a>) is the development of extremely inexpensive (e.g. $1000) systems for implementing nanobiotechnologies/molecular biology, especially for medical diagnostic applications.</div><br />
</td><br />
</tr><br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><br />
<br />
<strong>Robert Campbell</strong><br />
<br><br />
Department of Chemistry<br />
<br><br />
robert.e.campbell@ualberta.ca<br />
<img src="http://www.chem.ualberta.ca/faculty_staff/faculty/facultypics/campbell.jpg" width=200 height=274.19355 ALIGN="LEFT" hspace="20"></a><br />
</td><br />
<br />
<td><br />
<div align="justify" ><br />
Robert E. Campbell is an associate professor and Canada Research Chair in Bioanalytical Chemistry in the Department of Chemistry of the University of Alberta. Research in his laboratory, (<a href="http://www.chem.ualberta.ca/~campbell/">the Campbell Research Group</a>), is focused on protein engineering and the development of new fluorescent protein variants for construction of FRET-based biosensors.<br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><br />
<strong>Linda Reha-Krantz</strong><br />
<br><br />
Department of Biological Sciences<br />
<br><br />
Linda.Reha-Krantz@ualberta.ca<br />
<img src="http://www.biology.ualberta.ca/faculty/bio-187/uploads/images/reha_krantz.jpg" width=200 height=300 ALIGN="LEFT" hspace="20"></a><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</tr></td><br />
<br />
<tr><td><br />
<strong>Tracy Raivio</strong><br />
<br><br />
Department of Biological Sciences<br />
<br><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</tr></td><br />
<br />
<tr><td><br />
<strong>Jon Dennis</strong><br />
<br><br />
Department of Biological Sciences<br />
<br><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</tr></td><br />
<br />
<br />
<br />
<br />
</table><br />
<br />
</div><br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
</table><br />
</html></div>Mitch phttp://2009.igem.org/Team:Alberta/TeamTeam:Alberta/Team2009-10-20T21:49:10Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/Template3}}<br />
<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#e1e1e1; margin:0 5px;}<br />
.b2f {height:1px; background:#e1e1e1; margin:0 3px;}<br />
.b3f {height:1px; background:#e1e1e1; margin:0 2px;}<br />
.b4f {height:2px; background:#e1e1e1; margin:0 1px;}<br />
.content {background: #e1e1e1;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=60% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Recoli"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>Project BioBytes</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<br />
<center><br />
<img src="https://static.igem.org/mediawiki/2009/6/67/UofA_iGEM2009_IMG_5009.JPG" width="320" height="407"><br />
</center><br />
<br />
<br> <br />
<br />
<table><br />
<br />
<tr><br />
<td><br />
<div align="left"><strong>Kalon Armstrong</strong> <br><br />
<em>Molecular Genetics</em> <br><br />
</div><br />
<td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/3/34/UofA09_team_Kalon_Armstrong.jpg" hspace="20"><br />
</td><br />
<td><br />
<div align="justify" >I've recently completed my BSc in Molecular Genetics and will be entering the Engineering program in the fall of 2009. Most of my growing-up took place in the small town of Cochrane, Alberta. My ultimate goals consist of working in the Biotechnology or health care industry. While my interest in music, movies, snowboarding, and hanging out with friends may seem stereotypical on the surface, they feel unique in their own right and keep me busy most of the time. This will be my first year on the U of A iGEM team and I am excited to help take the competition to a new level. I think that iGEM will be a great experience because it demands innovation and collaboration on levels rarely seen in undergraduate programs.</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td colspan = "2"><br />
<br><br />
<br><br />
<hr style=width:66% align=left><br />
</td><br />
<tr><br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Eric Bennett</strong><br><br />
<em>Electrical Biomedical Engineering</em><br><br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/7/7a/UofA09_team_Eric_Bennett.jpg" hspace="20"><br />
</td><br />
<br />
<td><br />
<div align="justify" >I am entering my final year of engineering at the U of A. After graduation, I hope to do research either with a biotechnology company or in graduate level studies. I think that iGEM is a great way to gain valuable experience and is an effective way of accelerating the field of synthetic biology. My interests include brain-machine interfacing, genetic engineering, and robotic control systems. My hobbies include playing guitar, video games, the occasional sport, reading, and fixing my car.</div><br />
</td><br />
</tr> <br />
<br />
<tr><br />
<td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</tr><br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Max Buchko</strong><br><br />
<em>Honors Biochemistry</em><br><br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/e/e1/UofA09_team_Max_Buchko.jpg" ALIGN="LEFT" hspace="20"><br />
</td><br />
<br />
<td><br />
<div align="justify" style="padding-right:20px">I am in my third year of Honors Biochemistry and wish to pursue a career in medicine. In my time away from the lab I enjoy a wide variety of sports including soccer, boxing, and rifle silhouette shooting. I have also been known to strum a chord or two at an intolerable volume to the annoyance of the people living upstairs.<br />
I hope for the best with iGEM at MIT in 2009. This competition is a means of proving yourself at an exemplary level amongst many of the top international minds, and it is this challenge that I look forward to the most.</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td colspan = "2"><br />
<br><br />
<br><br />
<hr style=width:66% align=left><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Oscar Cortes</strong><br><br />
<em>Bsc. Specialization molecular genetics</em><br><br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/7/70/UofA09_team_Oscar_Cortes.jpg" ALIGN="LEFT" hspace="20"><br />
</td><br />
<td><br />
<div align="justify" style="padding-right:20px">My future plans include entering into a Masters program in Medical Genetics or Human Genetics, and pursuing this discipline towards a PhD. I enjoy reading books about the human genome and advancements in stem cell research. In my spare time I like to play a variety of sports that I am not necessarily good at: soccer, softball, and dodge ball. I have also been known to pound on a drum set from time to time without any rhythm whatsoever. I see iGEM as an extraordinary opportunity for me to be exposed to real life research, in which my knowledge of molecular genetics will be challenged, and will help me further my understanding of Synthetic Biology.<br />
"Man with all his noble qualities still bears in his bodily frame the indelible stamp of his lowly origin"- Charles Darwin</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</td><br />
</tr> <br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Anh Dao</strong><br><br />
<em>Biological Sciences and Chemistry</em><br> <br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/c/cf/UofA09_team_Ahn_Dao.jpg" ALIGN="LEFT" hspace="20"><br />
</td><br />
<br />
<td><br />
<div align="justify" style="padding-right:20px">I am interested in a research career after I finish my degree. I am leaning towards the field of Microbiology to study the many micro-organisms that have not yet been discovered. However, I may enroll in graduate studies after my undergraduate degree to expand my knowledge and gain more experience in the laboratory. Being in a competitive team and atmosphere is a motivating and exciting opportunity that I do not want to miss out on. By creating the smallest artificial E. coli genome we can extend future research. We are attempting to understand and standardize the E. coli genome so that this methodology can be applied to more complex model organisms.</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Uchechukwu Davidson</strong><br><br />
<em>Honors biochemistry</em><br><br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/3/3c/UofA09_team_Uche_Davidson.jpg" ALIGN="LEFT" hspace="20"><br />
</td><br />
<td><br />
<div align="justify" style="padding-right:20px">I am in my third year of Biochemistry at the University of Alberta. I wish to pursue a career in medicine after my degree. I enjoy sports, movies, and music at my time away from my studies. The iGEM provides an opportunity to experience creativity, innovation, and ingenuity which are sometimes absent at the undergraduate level of science.</div><br />
</td><br />
</tr><br />
<br />
<tr> <br />
<td colspan = "2"><br />
<br><br />
<br><br />
<br><br />
<hr style=width:66% align=left><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Youness Elkhalidy</strong><br><br />
<em>Honors immunology and infection</em><br><br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/f/fe/UofAiGEM2009_Youness_IMG_5049.JPG" width="200" height="300" ALIGN="LEFT" hspace="20"><br />
</td><br />
<td><br />
I am a first year student at the University of Alberta. I hope to enter medical school in the near future. I am currently taking part in mitotic-spindle regulation research from a genetics perspective. I have a passion for science and enjoy sports such as basketball and soccer. iGEM is an great learning experience not only in the cutting-edge field of Synthetic Biology but also in leadership and business management. I will enjoy taking part in research that combines many fields of science as well as socialize with many of my team members who share common interests.<br />
</td><br />
</tr><br />
<br />
<tr><br />
<td colspan = "2"><br />
<br />
<hr style=width:66% align=left><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Justin Fedor</strong><br><br />
<em>Honours Biochemistry</em><br><br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td><br />
<img src="https://static.igem.org/mediawiki/2009/7/71/UofA09_team_Justin_Fedor.jpg" ALIGN="LEFT" hspace="20"><br />
</td><br />
<br />
<td><br />
<div align="justify" style="padding-right:20px">I have completed my undergraduate degree in Biochemistry this year and will be starting my Ph.D. program in September. I play piano and am attempting to learn the cello. My nerdy tendencies are obvious when I say that I like Star Trek TNG but not DS9. Last summer I worked in the biomedical research lab of Dr. Larry Unsworth of the National Institute for Nanotechnology (NINT), which has further piqued my interest in the field of nanotechnology. In the hopefully not too distant future I plan on becoming a researcher studying the mechanisms of membrane bound enzymes, particularly oxidoreductases.</div><br />
</td><br />
</tr><br />
<br />
<tr><td colspan = "2"> <br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><br />
<td><br />
<div align="left"><br />
<strong>Jason Gardiner</strong> <br><br />
<em>BSc. Specialization in Botany</em><br><br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/2/2b/UofA09_team_Jason_Gardiner.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">Jason is a veteran of iGEM 2007, where his team "The Butanerds" won first place in the Energy track. Jason is currently in his fourth year of his Bachelor degree specializing in Botany. His hobbies include Softball, Beach volleyball, soccer, as well as playing the guitar and fiddling with the iGEM 2009 Wiki. Jason has applied to continue his education in graduate studies at the University of Alberta in 2010. His degree in Botany focuses mostly on the molecular side of plants and he hopes to use this knowledge applying Synthetic Biology to plants. One day he hopes to solve all of the worlds problems using plants and Synthetic Biology.</div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><br />
<div align="left"><br />
<strong>Erin Garside</strong> <br><br />
<em>Biological Sciences</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/6/60/UofA09_team_Erin_Garside.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">I completed my BSc. degree this year and plan to do graduate work in biochemistry. After that - who knows? I think iGEM will be a great experience, and that it's about time Synthetic Biology really took off. In my spare time I like to read, play computer games (especially the Sims) and raise cats.</div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Boris Henriquez</strong><br><br />
<em>Biological Sciences</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/e/ec/UofA09_team_Boris_Heneriquez.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I have just completed the third year of my BSc. degree. My favorite classes so far have been in the fields of Biochemistry and Genetics. I am most interested in human development and embryogenesis and hope to learn more about these topics in my fourth year. My ultimate goal is to have a career in the medical industry as both a physician and as a researcher. When I'm not busy with school or work my usual activities are pretty standard. I like to hang out with friends and family and I crave the outdoors. This is my first year with iGEM and I'm really excited to learn more about Synthetic Biology. </div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<br><br />
<br><br />
<br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Stephen Jahns</strong><br><br />
<em>Molecular Genetics</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/a/a8/Alberta_steve.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I am in my second year at the University of Alberta. After taking a year of engineering, I decided to transfer to molecular genetics in order to fulfill my childhood desire of creating an army of giant bat-rabbits. I plan on going on through to graduate school to earn a PhD in order to reach my goals. Also, health research could be pretty interesting too. In my spare time I like to run around, ride my bike, play music, cook delicious food, and pretty much all the other fun things kids are doing these days. In my first year I helped to build a car with the U of A's Formula SAE fabrication team. I had a good time building stuff out of metal, and this year I know that I'm going to have a blast building (or at least attempting to build) a novel organism out of chunks of DNA. I have a lot to learn but I know it will be worth it. </div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<br><br />
<br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<br />
<tr><td><div align="left"><br />
<strong>Eric Leung</strong> <br><br />
<em>Honors Pharmacology</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/a/a1/UofA09_team_Eric_Leung.jpg" ALIGN="LEFT" hspace="20"><br />
<br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">I am currently in my fourth and final year of an Honors Pharmacology degree. What I do after this degree is up in the air but it is definitely something in the health sciences. I hope I can become a professional baker in my spare time. Traveling to Europe, Mexico, and Japan is also in the books somewhere along the way.</div><br />
<br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<br />
<tr><td><div align="left"><br />
<strong>David Lloyd</strong><br> <br />
<em> Biochemistry</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/6/6f/UofA09_team_David_Lloyd.jpg" ALIGN="LEFT" hspace="20"><br />
<br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">I am a fourth year student, studying at the University of Alberta. While Biochemistry is my major, I have many interests including Genetics, Immunology, Cell Biology, Bioinformatics, and Microbiology. In my spare time I can be found playing sports like soccer, volleyball, or squash, as well as playing piano, listening to music, and playing video games. In the future, I hope to continue into a graduate program in Biochemistry, Cell Biology, Synthetic Biology, or in another field. iGEM is of great interest to myself because of its application to the future of Synthetic Biology. It is an awesome challenge which will hopefully help me to cultivate myself.</div><br />
<br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Enoch Ng</strong><br><br />
<em>Biological Sciences/Business</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/7/7c/UofA09_team_Enoch_Ng.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I am entering my fourth year of studies at the University of Alberta, and am looking forward to the challenge that iGEM provides. In the future I hope to travel across the world and meet people ranging from Meshaal to Obama and remain a glorified generalist. </div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Emera Nguyen</strong><br><br />
<em>BSc. Biological Sciences/Economics</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/5/52/UofA09_team_Emera_Nguyen.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I am a BSc student entering my fourth year in a Biological Sciences major and Economics minor. This year will be my second year on an iGEM team, and my first year representing the U of A team. In my downtime, things I enjoy include spending quality time with friends and family and traveling to lesser-known parts of the world. One year's experience later, the iGEM competition still impresses me with the unique opportunities it provides for student growth. This research competition is one of the highlights of my undergraduate experience.</div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr> <br />
<br />
<br />
<tr><td><div align="left"><br />
<strong>Mitch Paquette</strong><br><br />
<em>BSc. Honors Molecular Genetics</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/b/bf/Mitch5012_uofa.igem09.jpg" ALIGN="LEFT" hspace="20"><br />
<br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">I have transfered into the honors program in Molecular Genetics from a BSc General program in physical science. After my degree is completed I plan to apply to graduate studies in Molecular Biology, after which I hope to work in research or academia. What excites me most about iGEM is the opportunity to gain research experience in such a new field.</div><br />
</td></tr> <br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Amber Paul</strong><br><br />
<em>BSc. Specialization Immunology and Infection</em><br><br />
</div></td></tr><br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/3/3b/UofA_Team_Amber.jpg" width="200" height="300" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I plan to find the cure for cancer. Seriously. Will be attending graduate school to earn a PhD following my undergraduate degree. I would like to live somewhere warm that allows me to do research, preferably Maui. iGEM is a developmental forefront to Synthetic Biology, something that I love to be part of. Its challenging and fun. To be able to determine the minimal genes for a simple prokaryote, we provide a scaffold to future research on larger, more complex cells such as cancerous tissues, simple eukaryotes, etc.</div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Julia Pon</strong><br><br />
<em>Honors Molecular Genetics</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/3/30/UofA09_team_Julia_Pon.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">My future plans include pursuing PhD in Medical Genetics. I've worked in biological research labs for the past two summers, with last summer spent at the German Cancer Research Institute in Heidelberg, Germany. I'll be working in both the iGEM lab and a Medical Genetics research lab during summer 2009. My heritage is a mixture of English, Chinese, and Danish. iGEM helps move biology to a streamlined, standardized, abstracted process that opens the door to interdisciplinary collaborations and new applications. I'm excited about the many applications of Synthetic Biology and am enjoying the student-directed and team-oriented nature of iGEM.</div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<br />
<tr><td><div align="left"> <br />
<strong>Alina Ponomarev</strong><br><br />
<em>Biological Sciences</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/e/e1/UofA2009iGEM_Alina_IMG_5023.JPG" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I am currently a second year student in Biological Sciences and I am considering transferring into the Immunology and Infection program for my third year. I joined iGEM at the end of my first year and can honestly say that it has been an unbeatable learning experience. It is very exciting to think that the progress we have made can positively impact synthetic biology and society in general. In the future, my career goal is to become a Pediatrician. However, after doing labwork at the iGEM lab this summer, I have begun to consider a career in medical research. </div> <br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Kelly Robinson</strong><br><br />
<em>Hon. Biochemistry</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/9/92/UofA09_team_Kelly_Robinson.png" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">I completed my BSc. in Biochemistry this year and have enrolled in the University of Alberta's Chemical Engineering program in hopes of fusing the fundamental science of biological machines with the entrepreneurial world of engineering. I got involved in iGEM a couple of years ago when I went to a presentation put on by one of the past teams. I ended up applying in the middle of the night on the last day of registration. That said, it has been a great experience ever since. I'm glad that this year I get the opportunity to do some full time work and really get into the project. Overall, iGEM has a lot to offer anyone who gets involved (including incorrigible advisors: you know who you are!).</div><br />
<br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr> <br />
<br />
<tr><td><div align="left"><br />
<strong>James Rodway</strong><br><br />
<em>Electrical Engineering</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/a/a5/UofA09_team_James_Rodway.jpg" ALIGN="LEFT" hspace="20"><br />
<br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">I am currently finishing up an Electrical Engineering degree, and am aiming to do some graduate work in more of the computer area, specifically modeling. I've worked at a few cool places during co-op work terms and have done things like environmental monitoring and a whole mess of smaller projects during my time in the 'cube farm'. At the University I've been involved with the ARVP and iGEM, as best as time has allowed me.<br />
I haven't really had too much time for hobbies in the last while, but when I did I played more video games, the guitar, and I actually read books for entertainment value. During my time on last year's iGEM team I found that it was a pretty cool multidisciplinary project, which drew in a few different disciplines that I have never really interacted with at all previously. It was very impressive to see what we, and all the iGEM teams, had accomplished by the end of last year's competition.</div><br />
<br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Andy Spencer</strong><br><br />
<em>Honours Biochemistry</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/c/ca/UofA09_team_Andy_Spencer.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">In 5 years I see myself on one of two different paths. I will start applying to medical colleges this summer. Alternatively, I am interested in discovering novel treatments for cancer, and am considering a career in Research Oncology. I grew up in the Okanagan, British Columbia, and love spending my summers at the beach . In my spare time I enjoy playing squash and bouldering. iGEM to me represents a fundamental value of science as a discipline; that people can from diverse backgrounds with a common interest and curiosity can work as a cohesive team to overcome barriers and problems. I am excited to meet iGEM participants from around the world, and to visit MIT in November.</div><br />
<br />
</td></tr> <br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Jonathan Tam</strong><br><br />
<em>Honors Cell Biotechnology</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/e/e2/UofA09_team_Jonathan_Tam.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I have completed my Bachelor's degree in Honors Cell Biotechnology and intend to pursue a medical degree in Germany in the near future. I am very interested in the fields of Molecular Biology and Immunology. For the last two years, I have been keeping myself busy studying the development of macrophages in the lab of Dr. Daniel Barreda. Outside of the lab, I am an avid photographer and downhill mountain biker. The iGEM competition accelerates the development of Synthetic Biology as a field. To be a part of our University of Alberta team is an excellent opportunity for collaboration and advancement.</div><br />
</td></tr> <br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><div align="left"><br />
<strong>Jennifer Yau</strong><br><br />
<em>Honors Biochemistry</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/a/ac/UofA09_team_Jennifer_Yau.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<div align="justify" style="padding-right:20px">I am currently in the second year of my undergraduate program with the ultimate goal of attending graduate school to pursue a research career in Geriatric Medicine. Additional science related activities of mine include my interest in astronomy and promoting science to elementary students through volunteer work. Otherwise, I dedicate the majority of my free time to the arts of chainmaille, knitting, and jazz/classical piano. This is my first year on iGEM and I am ecstatic to be partaking in a project that requires such diverse disciplinary backgrounds. Not only will this be a great learning experience in so many perspectives, but also an opportunity to contribute significant ideas to the ongoing research in Synthetic Biology</div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr> <br />
<br />
<tr><td><div align="left"><br />
<strong>Zach Wiltshire</strong> <br><br />
<em>BSc. Specialization in Cell Biology</em><br><br />
</div></td></tr><br />
<br />
<tr><td><br />
<img src="https://static.igem.org/mediawiki/2009/1/19/UofA09_team_Zach_Wiltshire.jpg" ALIGN="LEFT" hspace="20"><br />
</td><td><br />
<br />
<div align="justify" style="padding-right:20px">I am entering into my fourth year in the Cell Biology program at the U of A and have already experienced iGEM once before as a member of the 2008 National Institute for Nanotechnology team. Through my degree I have found myself to have interests ranging from immunology to both eukaryotic & prokaryotic cellular anatomy & physiology. I also happen to be very fond of molecules that fluoresce. My experience with iGEM last year opened my eyes to the possibilities which engineering biological systems can offer. Over the course of the 2009 competition I hope to have just as many opportunities to learn as I did last year.</div><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<tr><td><br />
<a name="Team_Ad"></a><br />
<strong>The Supervisors</strong><br />
</tr><td><br />
<br />
<tr><td><br />
<strong>Mike Ellison</strong><br />
<img src="https://static.igem.org/mediawiki/2009/d/d5/UofA09_teamad_Mike_Ellison.jpg" ALIGN="LEFT" hspace="20"><br />
</td></tr><br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><br />
<br />
<strong>Doug Ridgway</strong><br />
<img src="https://static.igem.org/mediawiki/2009/c/cc/UofA09_teamad_Douglas_Ridgway.jpg" ALIGN="LEFT" hspace="20"></a><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><br />
<strong>James Maclagan</strong><br />
<img src="https://static.igem.org/mediawiki/2009/a/ab/UofAigem2009_JamesMIMG_4975.JPG" ALIGN="LEFT" hspace="20"></a><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</tr></td><br />
<tr><td><br />
<a name="Fac_Con"></a><br />
<strong>Faculty Consultants</strong><br />
</tr><td><br />
<br />
<tr><td><br />
<strong>Chris Backhouse</strong><br />
<br><br />
Department of Electrical Engineering<br />
<br><br />
christopher.backhouse@ualberta.ca<br />
<img src="http://www.ece.ualberta.ca/~chrisb/Graphics/Backhouse.jpg" ALIGN="LEFT" hspace="20"><br />
</td><br />
<br />
<td><br />
<div align="justify" ><br />
Nanobiotechnologies give us the ability to manipulate and sense at the level of individual molecules, with a tremendous potential impact on both human health and the economy. To a large extent, this potential is likely to be realised through the development of Lab on Chip (LOC) technologies. Although the LOC technologies are powerful, the complexity of the infrastructure required to support LOC operation has hindered the widespread adoption of LOC methods in life science applications. A central theme in the work of the Backhouse lab (<a href="http://www.ece.ualberta.ca/~aml/">the Applied Miniaturisation Lab, AML</a>) is the development of extremely inexpensive (e.g. $1000) systems for implementing nanobiotechnologies/molecular biology, especially for medical diagnostic applications.</div><br />
</td><br />
</tr><br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><br />
<br />
<strong>Robert Campbell</strong><br />
<br><br />
Department of Chemistry<br />
<br><br />
robert.e.campbell@ualberta.ca<br />
<img src="http://www.chem.ualberta.ca/faculty_staff/faculty/facultypics/campbell.jpg" width=200 height=274.19355 ALIGN="LEFT" hspace="20"></a><br />
</td><br />
<br />
<td><br />
<div align="justify" ><br />
Robert E. Campbell is an associate professor and Canada Research Chair in Bioanalytical Chemistry in the Department of Chemistry of the University of Alberta. Research in his laboratory, (<a href="http://www.chem.ualberta.ca/~campbell/">the Campbell Research Group</a>), is focused on protein engineering and the development of new fluorescent protein variants for construction of FRET-based biosensors.<br />
</div><br />
</td><br />
</tr><br />
<br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</td></tr><br />
<br />
<tr><td><br />
<strong>Linda Reha-Krantz</strong><br />
<br><br />
Department of Biological Sciences<br />
<br><br />
Linda.Reha-Krantz@ualberta.ca<br />
<img src="http://www.biology.ualberta.ca/faculty/bio-187/uploads/images/reha_krantz.jpg" width=200 height=300 ALIGN="LEFT" hspace="20"></a><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</tr></td><br />
<br />
<tr><td><br />
<strong>Tracy Raivio</strong><br />
<br><br />
Department of Biological Sciences<br />
<br><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</tr></td><br />
<br />
<tr><td><br />
<strong>Jon Dennis</strong><br />
<br><br />
Department of Biological Sciences<br />
<br><br />
</td></tr><br />
<br />
<tr><td colspan = "2"><br />
<br><br />
<hr style=width:66% align=left><br />
</tr></td><br />
<br />
<br />
<br />
<br />
</table><br />
<br />
</div><br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
</table><br />
</html></div>Mitch phttp://2009.igem.org/File:Mitch5012_uofa.igem09.JPGFile:Mitch5012 uofa.igem09.JPG2009-10-20T21:46:18Z<p>Mitch p: </p>
<hr />
<div></div>Mitch phttp://2009.igem.org/Team:Alberta/MedalRequirementsTeam:Alberta/MedalRequirements2009-10-20T15:57:31Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#e1e1e1; margin:0 5px;}<br />
.b2f {height:1px; background:#e1e1e1; margin:0 3px;}<br />
.b3f {height:1px; background:#e1e1e1; margin:0 2px;}<br />
.b4f {height:2px; background:#e1e1e1; margin:0 1px;}<br />
.content {background: #e1e1e1;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>Medals and Area Prize Deliverables</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<h3> Bronze: </h3><br />
<p><b>Design and document Biobrick parts:</b> We’ve entered over 400 Biobrick parts in the Registry of Standard Biological Parts. See the Parts registry link the deliverables tab for a complete list. In summary, we’ve submitted: </p><br />
<li>2 backbone plasmids</li><br />
<li>12 promoters in pAB or pBA </li><br />
<li>2 terminators in pAB or pBA</li><br />
<li>12 reporters, selectable markers and genes in pAB or pBA </li><br />
<li>2 primers for sequencing out of pAB and pBA </li><br />
<li>4 USER primers for the Biobyte assembly method</li><br />
<li>188 primer pairs for essential E.coli genes</li><br />
<br />
<p>All submitted parts have been verified as the correct size using test digests and gel electrophoresis, and 28 parts have been sequenced to verify correct insertion in pAB or pBA. Sequencing files are posted on the Parts Registry as advanced sequence analyses.</p><br />
<br />
<b>Submit DNA:</b><br />
<p>We’ve submitted DNA for 26 parts, including 24 parts in pAB or pBA, and both the pAB and pBA plasmids. </p><br />
<br />
<h3>Silver:</h3><br />
<b>Demonstrate that at least one new part works as expected: </b><br />
<p>All 188 pairs of primers for essential genes submitted to the parts registry have been tested in PCR and shown to give a product of the correct size. pAB/BA primers for sequencing inserts in pAB/BA also worked as intended when used to verify part insertion. </p><br />
<br />
<b>Characterize the operation of at least one new biobrick part and enter this info: </b><br />
<p>The pAB and pBA plasmids and USER primers have been extensively tested in order to optimize the Biobytes assembly method, as documented in the DNA assembly section of this wiki. We’ve also tested several components in pAB and pBA using the Biobytes assembly method, and verified by gel electrophoresis and sequencing that they assemble properly. Moreover, we’ve characterized the behavior of parts for Biobyte assembly in a microfluidic chip, demonstrating successful construct production on a micro-scale.</p><br />
<br />
<h3>Gold:</h3> <br />
<p>Only one of the following criteria must be met for Gold. However, our team has met all four possible criteria:</p><br />
<br />
<b>Characterize or improve an existing biobrick part or device:</b><br />
<p>The GFP and RFP parts we have submitted in pAB and pBA are derived from preexisting biobrick parts. By adapting these parts to the Biobytes plasmids, we allow them to be used in the Biobytes assembly method, expanding their usefulness. </p><br />
<br />
<p>Furthermore, we adapted the Anderson collection of promoters to the Biobytes method. We removed NheI and AvrII cut sites from the consensus promoter and added two nucleotides downstream of the -10 region to place an A as the +1 nucleotide. These promoters are currently being tested by cloning them upstream of a ribosome binding site-red fluorescent protein segment, and checking for red colonies. </p><br />
<br />
<b>Help another iGEM team: </b><br />
<p>We constructed a plasmid for the University of Lethbridge team. </p><br />
<br />
<b>Develop and document a new technical standard:</b> <br />
<p> We have written and submitted an RFC detailing how the biobytes assembly method can be used, and including detailed protocol for how to perform this assembly method.</p><br />
<br />
<b>Outline and detail a new approach to the issue of human practices:</b><br />
<p> As a novel approach to human practices, we worked with eight internationally competitive debaters to produce a debate about the ethical and societal implications of developing artificially engineered organisms. This debate was filmed and is posted and summarized on our wiki for the benefit of the public. The extensive experience of the debaters brings an expertise about policy issues not commonly seen among science and engineering students. The diverse background of the debaters allows a wide range of opinions to be reflected that could have been imagined by an iGEM team alone. The format of debate allows both sides of an issue equal opportunity, and requires the use of well-reasoned arguments and evidence. Moreover, debates are fast paced and engaging, capturing an audience’s attention. The debate is a valuable resource for educating synthetic biologists about public reactions, assessing public knowledge about genetic engineering, helping policy makers make well-reasoned decisions, and helping the public form their own opinion of synthetic biology. </p><br />
<br />
<p>Moreover, we reached out to high school, summer camps, libraries and colleges, doing demonstration debates about ethics and presentations on the science of synthetic biology. We’ve reached over 230 students already and are scheduled to reach over 400 students by January. We’ve developed and posted resources for teaching synthetic biology to a wide range of age groups. From the feedback we’ve collected and analyzed students almost unanimously feel learning about synthetic biology is valuable. </p><br />
<br />
<h3> Special Prizes:</h3><br />
<br />
<h3> Best Model:</h3> <br />
<p>We’ve developed a novel system for comprehensively analyzing the entirety of the known metabolic network of E. coli. Through this system, one can assess the net biomass production of an E. coli in which any combination of metabolic gene are present, and thus assess which combinations of pathways are necessary for cell survival. We used this model to predict the minimal set of metabolic genes required for E.coli survival, towards the goal of producing a minimal E. coli genome. This minimal gene list included 117 essential genes never previously identified as essential for a minimal genome. Moreover, a graphical user interface has been developed for exploring the results of our minimal metabolism modeling. </p><br />
<br />
<h3> Best New Standard and Best Foundational Advance: </h3><br />
<p> The Biobytes RFC outlines what is currently the only method for DNA assembly that is fast, modular, sequential and in vitro. </p><br />
<br />
<p>Synthetic biology needs more than minor modifications to existing evolutionary plans, as in the case of Craig Ventor’s efforts with mycoplasma. We’ve developed a method allowing genome design to be based almost exclusively on artificial design principles, such as maximizing modularity by grouping common pathway components and components with similar levels of expression. This degree of organism control would be a milestone marking Synthetic Biology as a mature field. It would allow us to rapidly test, optimize and correct design principles on a simple chassis that could be reliably modeled. </p><br />
<br />
<p>There are currently two alternatives for gene assembly. The first, Biobricks, is modular but slow. The second, rhe use of unique long sticky ends for each piece, is fast but non modular. </p><br />
<br />
<b>Biobytes is the only method that is fast, modular, sequential and in vitro:</b><br />
<p> <b>Fast: </b> The addition of each DNA segment takes only 20min, a roughly 200 fold increase in speed from traditional cloning. Moreover, we’ve demonstrated the Biobytes method works on microfluidic chips and is automatable both on both lab bench and microfluidic scales. </P><br />
<p> <b> Modular: </b> this allows standard parts such as the backbone primers and USER primers to be reused, greatly reducing expenses for large scale projects. Once parts are in pAB or pBA, they can be rapidly assembled in any order, allowing easy testing of alternative designs. </P><br />
<p><b>Sequential: </b> Biobytes allows tight control over the order of gene assembly. New DNA segments can add only to the unanchored end, and only with their complementary end. Using two different sets of complementary ends prevents concatamerization of parts before assembly. </P><br />
<p><b>In vitro: </b>Using an organism as an intermediate is time consuming and limits one’s ability to control and assess the changes they’re making. With current advances in transformation, genome sized constructs assembled in vitro can later be transformed into an organism. Biobytes allows the in vitro assembly. </P><br />
<br />
<p> Overall, the Biobytes method gives synthetic biology the tools to Understand and organize complexity, Standardize robust parts, Use modular strategies and Rapidly test rational designs and computational models. With biobytes we can start asking the most fundamental questions - To what extent do the rules of engineering hold true for biology? To what degree does life equal the sum of its parts? </P><br />
<br />
<br />
<br />
<br />
<br />
<font size="2"><br />
<P> ---- </p><br />
<br />
<br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/Team:Alberta/Project/RecombineeringTeam:Alberta/Project/Recombineering2009-09-17T23:14:04Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#ADED7C; margin:0 5px;}<br />
.b2f {height:1px; background:#ADED7C; margin:0 3px;}<br />
.b3f {height:1px; background:#ADED7C; margin:0 2px;}<br />
.b4f {height:2px; background:#ADED7C; margin:0 1px;}<br />
.content {background: #ADED7C;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>What is Recombineering?</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<P><br />
Recombineering refers to the strategic use of recombination <i>in vivo</i> in order to reach a defined goal. In the case of BioBytes, a method is required to target the final construct to insertion at a specific place on the <i>E. coli</i> chromosome.</p> <br />
<br />
<P><br />
To do this successfully, three components must be taken into account: <br />
</p><br />
<P> - There must be a system for targeting the construct to a specific site for insertion </p><br />
<P> - Activation of the recombination system must be under experimenter control </p><br />
<P> - It must be possible to select for and verify colonies in which the insertion was successful </p><br />
<br />
</font></div><br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="What is recombineering?"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>Targeting<h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<br />
<P>The BioBytes team has chosen to use a recombination system from bacteriophage lambda. Lambda Red recombinase specifically recombines on the ends of a linear fragment of DNA. If the ends of this fragment are homologous to two separate sites on the <i>E. coli</i> chomosome, the genetic material between these two homologous regions will be exchanged. This will be the basis for our targeting system.</p><br />
<P>The homologous regions must be a minimum of 50 base pairs in length for recombination to occur at a significant frequency. This can be achieved in different ways:</p><br />
<P>First 5' extensions corresponding to the homologous sequence can be added to any gene using PCR amplification. This would allow a PCR product to be targeted to a specific site for insertion. Because our constructs will be recircularized and grown (see <i>DNA assembly</i>), this would require us to PCR each plasmid construct seperately in order to add the homologous regions to the ends.</p><br />
<P>An alternative is to use the genes on either end of our construct as the homologous regions. As an example, we could first locate a region of genes which were deemed inessential through literature review and our Matlab modelling. This region would necessarily be flanked by an essential gene at either end. We would then assemble a plasmid containing these two essential genes. If the insertion is successful, we would be left with a chromosome without this region of inessential genes:</p><br />
<img src="https://static.igem.org/mediawiki/2009/2/2a/UofA_iGEM_09_RecombineFig1.jpg"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Targeting"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>Inducible Recombination System<h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<br />
<br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/Team:Alberta/Project/RecombineeringTeam:Alberta/Project/Recombineering2009-09-17T21:39:52Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#ADED7C; margin:0 5px;}<br />
.b2f {height:1px; background:#ADED7C; margin:0 3px;}<br />
.b3f {height:1px; background:#ADED7C; margin:0 2px;}<br />
.b4f {height:2px; background:#ADED7C; margin:0 1px;}<br />
.content {background: #ADED7C;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>What is Recombineering?</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<P><br />
Recombineering refers to the strategic use of recombination <i>in vivo</i> in order to reach a defined goal. In the case of BioBytes, a method is required to target the final construct to insertion at a specific place on the <i>E. coli</i> chromosome.</p> <br />
<br />
<P><br />
To do this successfully, three components must be taken into account: <br />
</p><br />
<P> - There must be a system for targeting the construct to a specific site for insertion </p><br />
<P> - Activation of the recombination system must be under experimenter control </p><br />
<P> - It must be possible to select for and verify colonies in which the insertion was successful </p><br />
<br />
</font></div><br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="What is recombineering?"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>Targeting<h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<br />
<P>The BioBytes team has chosen to use a recombination system from bacteriophage lambda. Lambda Red recombinase specifically recombines on the ends of a linear fragment of DNA. If the ends of this fragment are homologous to two separate sites on the <i>E. coli</i> chomosome, the genetic material between these two homologous regions will be exchanged. This will be the basis for our targeting system.</p><br />
<P>The homologous regions must be a minimum of 50 base pairs in length for recombination to occur at a significant frequency. This can be achieved in different ways:</p><br />
<P>First 5' extensions corresponding to the homologous sequence can be added to any gene using PCR amplification. This would allow a PCR product to be targeted to a specific site for insertion. Because our constructs will be recircularized and grown (see <i>DNA assembly</i>), this would require us to PCR each plasmid construct seperately in order to add the homologous regions to the ends.</p><br />
<P>An alternative is to use the genes on either end of our construct as the homologous regions. As an example, we could first locate a region of genes which were deemed inessential through literature review and our Matlab modelling. This region would necessarily be flanked by an essential gene at either end. We would then assemble a plasmid containing these two essential genes. If the insertion is successful, we would be left with a chromosome without this region of inessential genes:</p><br />
<img src="https://static.igem.org/mediawiki/2009/2/2a/UofA_iGEM_09_RecombineFig1.jpg"><br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/Team:Alberta/Project/RecombineeringTeam:Alberta/Project/Recombineering2009-09-17T21:39:24Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#ADED7C; margin:0 5px;}<br />
.b2f {height:1px; background:#ADED7C; margin:0 3px;}<br />
.b3f {height:1px; background:#ADED7C; margin:0 2px;}<br />
.b4f {height:2px; background:#ADED7C; margin:0 1px;}<br />
.content {background: #ADED7C;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>What is Recombineering?</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<P><br />
Recombineering refers to the strategic use of recombination <i>in vivo</i> in order to reach a defined goal. In the case of BioBytes, a method is required to target the final construct to insertion at a specific place on the <i>E. coli</i> chromosome.</p> <br />
<br />
<P><br />
To do this successfully, three components must be taken into account: <br />
</p><br />
<P> - There must be a system for targeting the construct to a specific site for insertion </p><br />
<P> - Activation of the recombination system must be under experimenter control </p><br />
<P> - It must be possible to select for and verify colonies in which the insertion was successful </p><br />
<br />
</font></div><br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="What is recombineering?"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>Targeting<h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<br />
<P>The BioBytes team has chosen to use a recombination system from bacteriophage lambda. Lambda Red recombinase specifically recombines on the ends of a linear fragment of DNA. If the ends of this fragment are homologous to two separate sites on the <i>E. coli</i> chomosome, the genetic material between these two homologous regions will be exchanged. This will be the basis for our targeting system.</p><br />
<P>The homologous regions must be a minimum of 50 base pairs in length for recombination to occur at a significant frequency. This can be achieved in different ways:</p><br />
<P>First 5' extensions corresponding to the homologous sequence can be added to any gene using PCR amplification. This would allow a PCR product to be targeted to a specific site for insertion. Because our constructs will be recircularized and grown (see <i>DNA assembly</i>), this would require us to PCR each plasmid construct seperately in order to add the homologous regions to the ends.</p><br />
<P>An alternative is to use the genes on either end of our construct as the homologous regions. As an example, we could first locate a region of genes which were deemed inessential through literature review and our Matlab modelling. This region would necessarily be flanked by an essential gene at either end. We would then assemble a plasmid containing these two essential genes. If the insertion is successful, we would be left with a chromosome without this region of inessential genes:</p><br />
<img src="https://static.igem.org/mediawiki/2009/2/2a/UofA_iGEM_09_RecombineFig1.jpeg"><br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/File:UofA_iGEM09_RecombineFig1.JPGFile:UofA iGEM09 RecombineFig1.JPG2009-09-17T21:38:36Z<p>Mitch p: </p>
<hr />
<div></div>Mitch phttp://2009.igem.org/Team:Alberta/Project/RecombineeringTeam:Alberta/Project/Recombineering2009-09-17T21:37:43Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#ADED7C; margin:0 5px;}<br />
.b2f {height:1px; background:#ADED7C; margin:0 3px;}<br />
.b3f {height:1px; background:#ADED7C; margin:0 2px;}<br />
.b4f {height:2px; background:#ADED7C; margin:0 1px;}<br />
.content {background: #ADED7C;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>What is Recombineering?</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<P><br />
Recombineering refers to the strategic use of recombination <i>in vivo</i> in order to reach a defined goal. In the case of BioBytes, a method is required to target the final construct to insertion at a specific place on the <i>E. coli</i> chromosome.</p> <br />
<br />
<P><br />
To do this successfully, three components must be taken into account: <br />
</p><br />
<P> - There must be a system for targeting the construct to a specific site for insertion </p><br />
<P> - Activation of the recombination system must be under experimenter control </p><br />
<P> - It must be possible to select for and verify colonies in which the insertion was successful </p><br />
<br />
</font></div><br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="What is recombineering?"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>Targeting<h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<br />
<P>The BioBytes team has chosen to use a recombination system from bacteriophage lambda. Lambda Red recombinase specifically recombines on the ends of a linear fragment of DNA. If the ends of this fragment are homologous to two separate sites on the <i>E. coli</i> chomosome, the genetic material between these two homologous regions will be exchanged. This will be the basis for our targeting system.</p><br />
<P>The homologous regions must be a minimum of 50 base pairs in length for recombination to occur at a significant frequency. This can be achieved in different ways:</p><br />
<P>First 5' extensions corresponding to the homologous sequence can be added to any gene using PCR amplification. This would allow a PCR product to be targeted to a specific site for insertion. Because our constructs will be recircularized and grown (see <i>DNA assembly</i>), this would require us to PCR each plasmid construct seperately in order to add the homologous regions to the ends.</p><br />
<P>An alternative is to use the genes on either end of our construct as the homologous regions. As an example, we could first locate a region of genes which were deemed inessential through literature review and our Matlab modelling. This region would necessarily be flanked by an essential gene at either end. We would then assemble a plasmid containing these two essential genes. If the insertion is successful, we would be left with a chromosome without this region of inessential genes:</p><br />
<img src="https://static.igem.org/mediawiki/2009/5/5e/UofA_iGEM_09_RecombineeringFig1.PNG"><br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/Team:Alberta/Project/RecombineeringTeam:Alberta/Project/Recombineering2009-09-17T21:37:32Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#ADED7C; margin:0 5px;}<br />
.b2f {height:1px; background:#ADED7C; margin:0 3px;}<br />
.b3f {height:1px; background:#ADED7C; margin:0 2px;}<br />
.b4f {height:2px; background:#ADED7C; margin:0 1px;}<br />
.content {background: #ADED7C;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>What is Recombineering?</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<P><br />
Recombineering refers to the strategic use of recombination <i>in vivo</i> in order to reach a defined goal. In the case of BioBytes, a method is required to target the final construct to insertion at a specific place on the <i>E. coli</i> chromosome.</p> <br />
<br />
<P><br />
To do this successfully, three components must be taken into account: <br />
</p><br />
<P> - There must be a system for targeting the construct to a specific site for insertion </p><br />
<P> - Activation of the recombination system must be under experimenter control </p><br />
<P> - It must be possible to select for and verify colonies in which the insertion was successful </p><br />
<br />
</font></div><br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="What is recombineering?"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>Targeting<h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<br />
<P>The BioBytes team has chosen to use a recombination system from bacteriophage lambda. Lambda Red recombinase specifically recombines on the ends of a linear fragment of DNA. If the ends of this fragment are homologous to two separate sites on the <i>E. coli</i> chomosome, the genetic material between these two homologous regions will be exchanged. This will be the basis for our targeting system.</p><br />
<P>The homologous regions must be a minimum of 50 base pairs in length for recombination to occur at a significant frequency. This can be achieved in different ways:</p><br />
<P>First 5' extensions corresponding to the homologous sequence can be added to any gene using PCR amplification. This would allow a PCR product to be targeted to a specific site for insertion. Because our constructs will be recircularized and grown (see <i>DNA assembly</i>), this would require us to PCR each plasmid construct seperately in order to add the homologous regions to the ends.</p><br />
<P>An alternative is to use the genes on either end of our construct as the homologous regions. As an example, we could first locate a region of genes which were deemed inessential through literature review and our Matlab modelling. This region would necessarily be flanked by an essential gene at either end. We would then assemble a plasmid containing these two essential genes. If the insertion is successful, we would be left with a chromosome without this region of inessential genes:</p><br />
<img src="https://static.igem.org/mediawiki/2009/5/5e/UofA_iGEM_09_RecombineeringFig1.PNG><br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/Team:Alberta/Project/RecombineeringTeam:Alberta/Project/Recombineering2009-09-17T21:37:19Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#ADED7C; margin:0 5px;}<br />
.b2f {height:1px; background:#ADED7C; margin:0 3px;}<br />
.b3f {height:1px; background:#ADED7C; margin:0 2px;}<br />
.b4f {height:2px; background:#ADED7C; margin:0 1px;}<br />
.content {background: #ADED7C;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>What is Recombineering?</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<P><br />
Recombineering refers to the strategic use of recombination <i>in vivo</i> in order to reach a defined goal. In the case of BioBytes, a method is required to target the final construct to insertion at a specific place on the <i>E. coli</i> chromosome.</p> <br />
<br />
<P><br />
To do this successfully, three components must be taken into account: <br />
</p><br />
<P> - There must be a system for targeting the construct to a specific site for insertion </p><br />
<P> - Activation of the recombination system must be under experimenter control </p><br />
<P> - It must be possible to select for and verify colonies in which the insertion was successful </p><br />
<br />
</font></div><br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="What is recombineering?"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>Targeting<h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<br />
<P>The BioBytes team has chosen to use a recombination system from bacteriophage lambda. Lambda Red recombinase specifically recombines on the ends of a linear fragment of DNA. If the ends of this fragment are homologous to two separate sites on the <i>E. coli</i> chomosome, the genetic material between these two homologous regions will be exchanged. This will be the basis for our targeting system.</p><br />
<P>The homologous regions must be a minimum of 50 base pairs in length for recombination to occur at a significant frequency. This can be achieved in different ways:</p><br />
<P>First 5' extensions corresponding to the homologous sequence can be added to any gene using PCR amplification. This would allow a PCR product to be targeted to a specific site for insertion. Because our constructs will be recircularized and grown (see <i>DNA assembly</i>), this would require us to PCR each plasmid construct seperately in order to add the homologous regions to the ends.</p><br />
<P>An alternative is to use the genes on either end of our construct as the homologous regions. As an example, we could first locate a region of genes which were deemed inessential through literature review and our Matlab modelling. This region would necessarily be flanked by an essential gene at either end. We would then assemble a plasmid containing these two essential genes. If the insertion is successful, we would be left with a chromosome without this region of inessential genes:</p><br />
<img src="https://static.igem.org/mediawiki/2009/5/5e/UofA_iGEM_09_RecombineeringFig1.PNG"><br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/Team:Alberta/Project/RecombineeringTeam:Alberta/Project/Recombineering2009-09-17T21:36:51Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#ADED7C; margin:0 5px;}<br />
.b2f {height:1px; background:#ADED7C; margin:0 3px;}<br />
.b3f {height:1px; background:#ADED7C; margin:0 2px;}<br />
.b4f {height:2px; background:#ADED7C; margin:0 1px;}<br />
.content {background: #ADED7C;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>What is Recombineering?</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<P><br />
Recombineering refers to the strategic use of recombination <i>in vivo</i> in order to reach a defined goal. In the case of BioBytes, a method is required to target the final construct to insertion at a specific place on the <i>E. coli</i> chromosome.</p> <br />
<br />
<P><br />
To do this successfully, three components must be taken into account: <br />
</p><br />
<P> - There must be a system for targeting the construct to a specific site for insertion </p><br />
<P> - Activation of the recombination system must be under experimenter control </p><br />
<P> - It must be possible to select for and verify colonies in which the insertion was successful </p><br />
<br />
</font></div><br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="What is recombineering?"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>Targeting<h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<br />
<P>The BioBytes team has chosen to use a recombination system from bacteriophage lambda. Lambda Red recombinase specifically recombines on the ends of a linear fragment of DNA. If the ends of this fragment are homologous to two separate sites on the <i>E. coli</i> chomosome, the genetic material between these two homologous regions will be exchanged. This will be the basis for our targeting system.</p><br />
<P>The homologous regions must be a minimum of 50 base pairs in length for recombination to occur at a significant frequency. This can be achieved in different ways:</p><br />
<P>First 5' extensions corresponding to the homologous sequence can be added to any gene using PCR amplification. This would allow a PCR product to be targeted to a specific site for insertion. Because our constructs will be recircularized and grown (see <i>DNA assembly</i>), this would require us to PCR each plasmid construct seperately in order to add the homologous regions to the ends.</p><br />
<P>An alternative is to use the genes on either end of our construct as the homologous regions. As an example, we could first locate a region of genes which were deemed inessential through literature review and our Matlab modelling. This region would necessarily be flanked by an essential gene at either end. We would then assemble a plasmid containing these two essential genes. If the insertion is successful, we would be left with a chromosome without this region of inessential genes:</p><br />
<img src="https://static.igem.org/mediawiki/2009/5/5e/UofA_iGEM_09_RecombineeringFig1.png"><br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/File:UofA_iGEM09_RecombineeringFig1.PNGFile:UofA iGEM09 RecombineeringFig1.PNG2009-09-17T21:33:12Z<p>Mitch p: </p>
<hr />
<div></div>Mitch phttp://2009.igem.org/Team:Alberta/Project/RecombineeringTeam:Alberta/Project/Recombineering2009-09-17T21:29:11Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#ADED7C; margin:0 5px;}<br />
.b2f {height:1px; background:#ADED7C; margin:0 3px;}<br />
.b3f {height:1px; background:#ADED7C; margin:0 2px;}<br />
.b4f {height:2px; background:#ADED7C; margin:0 1px;}<br />
.content {background: #ADED7C;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>What is Recombineering?</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<P><br />
Recombineering refers to the strategic use of recombination <i>in vivo</i> in order to reach a defined goal. In the case of BioBytes, a method is required to target the final construct to insertion at a specific place on the <i>E. coli</i> chromosome.</p> <br />
<br />
<P><br />
To do this successfully, three components must be taken into account: <br />
</p><br />
<P> - There must be a system for targeting the construct to a specific site for insertion </p><br />
<P> - Activation of the recombination system must be under experimenter control </p><br />
<P> - It must be possible to select for and verify colonies in which the insertion was successful </p><br />
<br />
</font></div><br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="What is recombineering?"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>Targeting<h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<br />
<P>The BioBytes team has chosen to use a recombination system from bacteriophage lambda. Lambda Red recombinase specifically recombines on the ends of a linear fragment of DNA. If the ends of this fragment are homologous to two separate sites on the <i>E. coli</i> chomosome, the genetic material between these two homologous regions will be exchanged. This will be the basis for our targeting system.</p><br />
<P>The homologous regions must be a minimum of 50 base pairs in length for recombination to occur at a significant frequency. This can be achieved in different ways:</p><br />
<P>First 5' extensions corresponding to the homologous sequence can be added to any gene using PCR amplification. This would allow a PCR product to be targeted to a specific site for insertion. Because our constructs will be recircularized and grown (see <i>DNA assembly</i>), this would require us to PCR each plasmid construct seperately in order to add the homologous regions to the ends.</p><br />
<P>An alternative is to use the genes on either end of our construct as the homologous regions. As an example, we could first locate a region of genes which were deemed inessential through literature review and our Matlab modelling. This region would necessarily be flanked by an essential gene at either end. We would then assemble a plasmid containing these two essential genes. If the insertion is successful, we would be left with a chromosome without this region of inessential genes:<br />
<br />
</table><br />
</div><br />
</HTML></div>Mitch phttp://2009.igem.org/Team:Alberta/Project/RecombineeringTeam:Alberta/Project/Recombineering2009-09-17T21:28:41Z<p>Mitch p: </p>
<hr />
<div>{{:Team:Alberta/TemplateSc}}<br />
<html><br />
<head><br />
<style type="text/css"><br />
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}<br />
.b1f {height:1px; background:#ADED7C; margin:0 5px;}<br />
.b2f {height:1px; background:#ADED7C; margin:0 3px;}<br />
.b3f {height:1px; background:#ADED7C; margin:0 2px;}<br />
.b4f {height:2px; background:#ADED7C; margin:0 1px;}<br />
.content {background: #ADED7C;}<br />
.content div {margin-left: 5px;}<br />
</style><br />
</head><br />
<br />
<div class="all"><br />
<div style="background:#FFFFFF"><br />
<br />
<!-- adjust table width, main background and padding between cells and edge of background --><br />
<br />
<br />
<table width=75% style="background:#FFFFFF; padding:2px;"><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="Outreach"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>What is Recombineering?</h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<P><br />
Recombineering refers to the strategic use of recombination <i>in vivo</i> in order to reach a defined goal. In the case of BioBytes, a method is required to target the final construct to insertion at a specific place on the <i>E. coli</i> chromosome.</p> <br />
<br />
<P><br />
To do this successfully, three components must be taken into account: <br />
</p><br />
<P> - There must be a system for targeting the construct to a specific site for insertion </p><br />
<P> - Activation of the recombination system must be under experimenter control </p><br />
<P> - It must be possible to select for and verify colonies in which the insertion was successful </p><br />
<br />
</font></div><br />
<br />
</div></div><br />
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b><br />
</td><br />
</tr><br />
<br />
<tr><br />
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;"><br />
<b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b><br />
<div class="What is recombineering?"><br />
<div style="height: 400; background:#FFFFFF; colorou line-height:100% padding: 3px 0px;"><br />
<h1>Targeting<h1><br />
<br />
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> --><br />
<div align="justify"><br />
<br />
<font size="2"><br />
<br />
<P>The BioBytes team has chosen to use a recombination system from bacteriophage lambda. Lambda Red recombinase specifically recombines on the ends of a linear fragment of DNA. If the ends of this fragment are homologous to two separate sites on the <i>E. coli</i> chomosome, the genetic material between these two homologous regions will be exchanged. This will be the basis for our targeting system.</p><br />
<P>The homologous regions must be a minimum of 50 base pairs in length for recombination to occur at a significant frequency. This can be achieved in different ways:</p><br />
<P>First 5' extensions corresponding to the homologous sequence can be added to any gene using PCR amplification. This would allow a PCR product to be targeted to a specific site for insertion. Because our constructs will be recircularized and grown (see <i>DNA assembly</i>), this would require us to PCR each plasmid construct seperately in order to add the homologous regions to the ends.</p><br />
<P>An alternative is to use the genes on either end of our construct as the homologous regions. As an example, we could first locate a region of genes which were deemed inessential through literature review and our Matlab modelling. This region would necessarily be flanked by an essential gene at either end. We would then assemble a plasmid containing these two essential genes. If the insertion is successful, we would be left with a chromosome without this region of inessential genes:</p><br />
<img src="https://static.igem.org/mediawiki/2009/a/ac/UofA_iGEM09_RecombFig1.png> <br />
</table><br />
</div><br />
</HTML></div>Mitch p