Team:Alberta

From 2009.igem.org

(Difference between revisions)
m
 
(41 intermediate revisions not shown)
Line 4: Line 4:
<style type="text/css">
<style type="text/css">
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}
.b1f, .b2f, .b3f, .b4f{font-size:1px; overflow:hidden; display:block;}
-
.b1f {height:1px; background:#ADED7C; margin:0 5px;}
+
.b1f {height:1px; background:#e1e1e1; margin:0 5px;}
-
.b2f {height:1px; background:#ADED7C; margin:0 3px;}
+
.b2f {height:1px; background:#e1e1e1; margin:0 3px;}
-
.b3f {height:1px; background:#ADED7C; margin:0 2px;}
+
.b3f {height:1px; background:#e1e1e1; margin:0 2px;}
-
.b4f {height:2px; background:#ADED7C; margin:0 1px;}
+
.b4f {height:2px; background:#e1e1e1; margin:0 1px;}
-
.content {background: #ADED7C;}
+
.content {background: #e1e1e1;}
.content div {margin-left: 5px;}
.content div {margin-left: 5px;}
</style>
</style>
Line 22: Line 22:
<tr>
<tr>
-
<td style="height: 400; padding-left: 10px; padding-right: 10px; padding-top: 11px;">
+
<td style="height: 200; padding-left: 10px; padding-right: 10px; padding-top: 11px;">
     <b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b>
     <b class="b1f"></b><b class="b2f"></b><b class="b3f"></b><b class="b4f"></b>
     <div class="Recoli">
     <div class="Recoli">
Line 28: Line 28:
     <h1>BioBytes</h1>
     <h1>BioBytes</h1>
-
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> -->
 
<div align="justify">
<div align="justify">
-
<font size="2">Team BioBytes is the University of Alberta's 2009 International Genetically Engineered Machines (iGEM) team. This year's iGEM project can be subdivided into two major efforts. The first and most important of which is the BioBytes chromosome assembly system. This system refers to a mechanism for rapid and reliable construction of plasmids (i.e.: artificial gene sets) in vitro. The second, the minimal genome project, refers to the ultimate goal of rapid and reliable DNA assembly, that is, the construction of an artificial <i>E. coli</i> chromosome. Furthermore, it includes the strategy of gene selection, arrangement, artificial chromosome insertion and the destruction of the host's chromosome.</P>
+
<font size="2">
 +
<p>Synthetic biology needs more than minor modifications to existing evolutionary plans. We’ve developed a method of gene assembly allowing complete genome re-design. The speed and automation of the Biobytes method makes possible the maximization of modularity on a grand scale. Imagine a synthetic genome 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. The Biobytes method of gene assembly allows us to efficiently test, optimize and correct genome scale design principles. </p>
 +
 
 +
<p>There are currently two alternatives for gene assembly. The first, BioBricks, is modular but slow. The second, the use of unique long sticky ends for each piece, is fast but non-modular. </p>
 +
<P>
 +
<b>BioBytes is the only method that is fast, modular, sequential and in vitro:</b></P>
 +
<ul>
 +
<li> <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 that the Biobytes method is automatable and can be performed on microfluidic chips.</li>
 +
<li> <b> Modular: </b> Our method allows standard parts such as the backbone plasmids 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.  </li>
 +
<li><b>Sequential: </b> Biobytes allows tight control over the order of gene assembly. New DNA segments can add only to the unanchored end, and in only one orientation. Moreover, using two different sets of complementary ends prevents concatamerization of parts before assembly.  </li>
 +
<li><b>In vitro: </b>Using an organism as an intermediate is time-consuming and limits one’s ability to control and assess the changes being made. For this reason, an in vitro method such as Biobytes is essential. Genome-sized constructs can be transformed into an organism after construction is complete. </li>
 +
</ul>
 +
<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>
 +
 
 +
<p align=right><a href="https://2009.igem.org/Team:Alberta/Project/assemblyoverview"> Click here for more...</a> </P>
 +
 
</font></div>
</font></div>
 +
       </div></div>
       </div></div>
Line 44: Line 59:
     <div class="Overview">
     <div class="Overview">
     <div style="height: 800; background:#FFFFFF; line-height:100% padding: 3px 0px;">
     <div style="height: 800; background:#FFFFFF; line-height:100% padding: 3px 0px;">
-
     <h2>The BioBytes Alternative </h2>
+
     <h2>Organism Design and The Minimal Genome Project</h2>
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> -->
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> -->
<div align="justify">
<div align="justify">
<font size="2">
<font size="2">
-
<p>BioBytes constitutes the University of Alberta’s contribution to iGEM 2009. It is a next-generation gene assembly system with the singular potential to accelerate the field toward the grand vision of the artificial cell. Unlike biobricks, genes produced in the BioByte format can be assembled in vitro rapidly, in any desired order, at great precision and yield. With cycle times approaching 15 minutes for the addition of each new gene, BioByte assembly rates exceed their biobrick counterpart by 200-fold. This level of improvement immediately opens the door towards the synthesis of simple chromosomes that can be tested and optimized at unprecedented speed. Finally, the BioByte assembly system requires a fraction of the equipment found in a conventional gene lab. This advantage combined with a "child-could-do-it" simplicity greatly extends its utility from the high school classroom to the workbench of the bio-process engineer. </p>
+
 
-
<p>A BioByte is a piece of DNA that encodes a specific type of cellular function or instruction. Each end of the DNA is distinct from the other so that they so that they can only be joined in a head-to-tail fashion as shown below. </p>
+
<p>The BioBytes gene assembly 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>
-
<p><strong>Figure 1.</strong></p>
+
 
-
<img src="https://static.igem.org/mediawiki/2009/a/a6/UofA_BBAlt_Figure1.png">
+
<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>
-
<p>Once correctly paired, Bytes are irreversibly linked together by a special enzyme called DNA ligase. It is the strength and accuracy of the head-to-tail interaction that accounts for BioByte’s superior yield, precision and speed relative to its BioBrick predecessor. </p>
+
 
-
<p>The hand-in-glove nature of the BioByte interaction illustrates how the orientation of each Byte is determined, but how can this approach be used to control the order by which Bytes are assembled? </p>
+
<p align=right><p align=right><a href="https://2009.igem.org/Team:Alberta/Project/Bioinformatics"> Click here for more...</a> </P>
-
<p><strong>Figure 2.</strong></p>
+
<P>
-
<img src="https://static.igem.org/mediawiki/2009/0/01/UofA_BBAlt_Figure2.png">
+
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. At the same time, the original host chromosome is displaced, effectively rebooting an <i>E.coli</i> cell with the synthetic chromosome. 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 ideal model organism</p>
-
<p>The problem of determining Byte order is shown in Figure 2. Here, the free ends on the existing two-byte chain means that the incoming third brick can be added randomly to either end resulting in the order [1-2-3] or [3-1-2]. BioBytes has overcome this problem by using a third type of end that can only be linked to an inert magnetic microsphere (Figure 3). By anchoring the first byte to the microsphere via this new end, only its free end is available for interaction. The chain therefore is constrained to grow in only one direction, away from its anchor. The microsphere design also fulfills another important function. Microspheres stick to magnets. Anchored chains can therefore be moved out of one reaction mixture leaving unlinked bytes behind, into a new reaction that containing new byte molecules needed for the next round of addition. </p>
+
 
-
<p><strong>Figure 3.</strong></p>
+
<p align=right><p align=right><a href="https://2009.igem.org/Team:Alberta/Project/Chromosome_Assembly"> Click here for more...</a> </P>
-
<img src="https://static.igem.org/mediawiki/2009/a/a3/UofA_BBAlt_Figure3a.png">
+
 
-
<img src="https://static.igem.org/mediawiki/2009/9/96/UofA_BBAlt_Figure3b.png">
+
-
<p>At this stage, the problem of Byte order and chain fidelity is not entirely solved. The method described above cannot exclude the possibility that multiple copies of a particular Byte become incorporated at a given step as shown in Figure 4 below. </p>
+
-
<p><strong>Figure 4.</strong></p>
+
-
<img src="https://static.igem.org/mediawiki/2009/4/46/UofA_BBAlt_Figure4.png">
+
-
<p>The problem arises because each byte that is incorporated into the chain does not enter the reaction mixture as single molecule but as a population of identical molecules that are as likely to interact with each other as the anchored chain (Figure 5). </p>
+
-
<p><strong>Figure 5.</strong></p>
+
-
<img src="https://static.igem.org/mediawiki/2009/8/88/UofA_BBAlt_Figure5.png">
+
-
<p>BioBytes solves the problem by constructing each Byte in two alternative forms, an “A” form and a “B” form. Each form has two incompatible ends. Therefore neither form can be linked to itself (Figure 6). The ends of each form are however, are compatible to each other, allowing for the alternating order of A and B forms in head-to-tail orientation. Adding Bytes to the growing chain by alternating the A an B forms assures that only one copy of each is added at each step </p>
+
-
<p><strong>Figure 6.</strong></p>
+
-
<img src="https://static.igem.org/mediawiki/2009/7/7e/UofA_BBAlt_Figure6.png">
+
-
<p>Upon completion of the desired product, chains are released from the microspheres by a chemical cleavage event that separates the anchor brick from its bound end, and are then introduced into living cells.</p>
+
-
<p>With its BioBytes approach, the Alberta team has recently demonstrated the accurate construction of chains composed of 10 Bytes over the course of 4 hours with no obvious limit to the final length that can be achieved. A key advantage to this approach is that it can be multiplexed for the production of many different chains simultaneously, that, can all be linked as SuperBytes by the same method to produce artificial chromosomes of unprecedented length.</p>
+
-
</font>
+
</font>
</font>
</div>
</div>
-
 
       </div></div>
       </div></div>
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b>
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b>
 +
     </td>
     </td>
   </tr>
   </tr>
-
 
<tr>
<tr>
<td style="height: 800; padding-left: 10px; padding-right: 10px; padding-top: 11px;">
<td style="height: 800; padding-left: 10px; padding-right: 10px; padding-top: 11px;">
Line 86: Line 87:
     <div class="Overview">
     <div class="Overview">
     <div style="height: 800; background:#FFFFFF; line-height:100% padding: 3px 0px;">
     <div style="height: 800; background:#FFFFFF; line-height:100% padding: 3px 0px;">
-
     <h2>The Minimal Genome Project</h2>
+
     <h2>Team Achievements</h2>
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> -->
<!-- <div align="justify" style="padding-left:20px; padding-right:20px"> -->
<div align="justify">
<div align="justify">
-
<font size="2"><P>The minimal <i>E. coli</i> genome has been a 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, in principle, preferred as a chassis for experimentation. To reduce the <i>E. coli</i> genome to roughly 10% its original size shows a great simplification of this model organism.</P>
+
<font size="2"><P> Through our efforts we have made the following accomplishments:</p>
-
<P>
+
<ul>
-
To create 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>. We then intend to remove the host chromosome by making it incapable of division. This allows only the artificial, inserted chromosome to propagate through multiple generations as the cells grow and divide. This is markedly different than 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>
+
<li>Developed the BioBytes Assembly Method and produced a proof of concept of the design
 +
<li>A kit of components has been added to the registry allowing for use of our system by anyone
 +
<li>Produced a series of modeling programs which can be used to determine the essential genes in any organism
 +
<li>185 essential genes have been amplified and their primers added to the registry
 +
<li>A robot has been developed demonstrating the potential of automation for BioBytes
 +
<li>Used microfluidics to show the biofabrication potential of our design
 +
<li>We have hosted debates involving synthetic biology
 +
<li>We have constructed and completed a series of presentations to discuss iGEM and promote knowledge of synthetic biology
 +
</ul>
 +
<p align=right><p align=right><a href="https://2009.igem.org/Team:Alberta/MedalRequirements"> Click here for more...</a> </P>
</font>
</font>
</div>
</div>
       </div></div>
       </div></div>
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b>
<b class="b4f"></b><b class="b3f"></b><b class="b2f"></b><b class="b1f"></b>
-
<center>
+
 
-
<table width="133" border="0" cellspacing="0" cellpadding="3"><tr><td align="center"><a href="http://www.website-hit-counters.com" target="_blank"><img src="http://www.website-hit-counters.com/cgi-bin/image.pl?URL=232857-4018" alt="" border="0" ></a></td></tr><tr><td align="center"><font style="font-family: Geneva, Arial, Helvetica, sans-serif; font-size: 9px; color: #330006; text-decoration: none;">Get a free <a href="http://www.website-hit-counters.com" target="_blank" style="font-family: Geneva, Arial, Helvetica, sans-serif; font-size: 9px; color: #555556; text-decoration: none;" title="html hit counter">html hit counter</a> here.</font></td></tr></table>
+
-
</center>
+
     </td>
     </td>
   </tr>
   </tr>
 +
 +
</table>
</table>
</div>
</div>
</HTML>
</HTML>

Latest revision as of 03:59, 22 October 2009

University of Alberta - BioBytes










































































































BioBytes

Synthetic biology needs more than minor modifications to existing evolutionary plans. We’ve developed a method of gene assembly allowing complete genome re-design. The speed and automation of the Biobytes method makes possible the maximization of modularity on a grand scale. Imagine a synthetic genome 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. The Biobytes method of gene assembly allows us to efficiently test, optimize and correct genome scale design principles.

There are currently two alternatives for gene assembly. The first, BioBricks, is modular but slow. The second, the use of unique long sticky ends for each piece, is fast but non-modular.

BioBytes is the only method that is fast, modular, sequential and in vitro:

  • Fast: The addition of each DNA segment takes only 20min, a roughly 200-fold increase in speed from traditional cloning. Moreover, we’ve demonstrated that the Biobytes method is automatable and can be performed on microfluidic chips.
  • Modular: Our method allows standard parts such as the backbone plasmids 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.
  • Sequential: Biobytes allows tight control over the order of gene assembly. New DNA segments can add only to the unanchored end, and in only one orientation. Moreover, using two different sets of complementary ends prevents concatamerization of parts before assembly.
  • In vitro: Using an organism as an intermediate is time-consuming and limits one’s ability to control and assess the changes being made. For this reason, an in vitro method such as Biobytes is essential. Genome-sized constructs can be transformed into an organism after construction is complete.

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?

Click here for more...

Organism Design and The Minimal Genome Project

The BioBytes gene assembly 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.

The minimal Escherichia coli genome has been the holy grail of biology for a number of years. E. coli 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 E. coli is ideally preferred as a chassis for experimentation. Reducing the E. coli genome to roughly 10% of its original size, demonstrates a great simplification of this model organism.

Click here for more...

To reconstruct such an organism, we plan on building an artificial E. coli chromosome using the BioBytes chromosome assembly system and inserting it into living E. coli cells. At the same time, the original host chromosome is displaced, effectively rebooting an E.coli cell with the synthetic chromosome. 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 ideal model organism

Click here for more...

Team Achievements

Through our efforts we have made the following accomplishments:

  • Developed the BioBytes Assembly Method and produced a proof of concept of the design
  • A kit of components has been added to the registry allowing for use of our system by anyone
  • Produced a series of modeling programs which can be used to determine the essential genes in any organism
  • 185 essential genes have been amplified and their primers added to the registry
  • A robot has been developed demonstrating the potential of automation for BioBytes
  • Used microfluidics to show the biofabrication potential of our design
  • We have hosted debates involving synthetic biology
  • We have constructed and completed a series of presentations to discuss iGEM and promote knowledge of synthetic biology

Click here for more...