Team:UC Davis/Parts
From 2009.igem.org
Line 49: | Line 49: | ||
<div style="text-align: left;"><big><big><b | <div style="text-align: left;"><big><big><b | ||
style="color: rgb(0, 0, 0); text-decoration: underline;">Parts:</b></big></big><b | style="color: rgb(0, 0, 0); text-decoration: underline;">Parts:</b></big></big><b | ||
- | style="color: rgb(0, 0, 0);"> <br> | + | style="color: rgb(0, 0, 0);"> <br> |
Parts related to | Parts related to | ||
- | secretion: | + | secretion: |
- | Parts related to pH sensor:<br> | + | |
+ | Parts related to pH | ||
+ | sensor:<br> | ||
</b> | </b> | ||
<table | <table | ||
Line 65: | Line 67: | ||
<td style="vertical-align: top;">Others:<br> | <td style="vertical-align: top;">Others:<br> | ||
</td> | </td> | ||
- | <td style="vertical-align: top;"> | + | <td style="vertical-align: top;"> New parts:<br> |
</td> | </td> | ||
<td style="vertical-align: top;"> | <td style="vertical-align: top;"> | ||
Promoters:<br> | Promoters:<br> | ||
</td> | </td> | ||
+ | <td style="vertical-align: top;">Proteins:</td> | ||
</tr> | </tr> | ||
<tr> | <tr> | ||
Line 95: | Line 98: | ||
<td style="vertical-align: top;"> | <td style="vertical-align: top;"> | ||
<ul> | <ul> | ||
- | <li><a href=" | + | <li><a href="#INPNCSS">INPNC + SS<br> |
- | <li><a | + | </a></li> |
- | href=" | + | <li><a href="#OmpAss">OmpA + SS<br> |
+ | </a></li> | ||
+ | <li><a href="#INPNC">INPNC </a><br> | ||
+ | </li> | ||
+ | <li><a href="#SS">SS</a><br> | ||
+ | </li> | ||
</ul> | </ul> | ||
</td> | </td> | ||
Line 109: | Line 117: | ||
</li> | </li> | ||
<li>I<a href="#impA">mpA promoter</a></li> | <li>I<a href="#impA">mpA promoter</a></li> | ||
+ | </ul> | ||
+ | </td> | ||
+ | <td style="vertical-align: top;"> | ||
+ | <ul> | ||
+ | <li><a href="https://2009.igem.org/Team:UC_Davis/ChvI1">ChvI</a></li> | ||
+ | <li><a | ||
+ | href="https://2009.igem.org/Team:UC_Davis/Project1/ChvG.html">ChvG</a></li> | ||
</ul> | </ul> | ||
</td> | </td> | ||
Line 119: | Line 134: | ||
</div> | </div> | ||
<hr style="width: 100%; height: 2px;"> | <hr style="width: 100%; height: 2px;"> | ||
+ | <p class="MsoNormal" | ||
+ | style="line-height: normal; font-weight: bold; text-align: center; text-decoration: underline;"><big><big>New | ||
+ | parts:</big></big></p> | ||
<p class="MsoNormal" style="line-height: normal;"><a name="INPNC"></a><b><span | <p class="MsoNormal" style="line-height: normal;"><a name="INPNC"></a><b><span | ||
style="font-size: 13.5pt; font-family: "Times New Roman","serif";">INPNC: | style="font-size: 13.5pt; font-family: "Times New Roman","serif";">INPNC: | ||
</span></b><span | </span></b><span | ||
style="font-size: 12pt; font-family: "Times New Roman","serif";">An | style="font-size: 12pt; font-family: "Times New Roman","serif";">An | ||
- | exogenous gene, | + | exogenous gene, Ice-nucleation |
- | Ice-nucleation protein (INP) from < | + | protein (INP) from <i>Pseudomonas syringae</i> was suggested to be |
- | syringae</ | + | used for |
- | be | + | display of foreign proteins on the surface of <i>E. coli. </i>(7). |
- | for display of foreign proteins on the surface of <i>E. coli</i>(7). | + | Furthermore, |
- | Furthermore, studies have shown that an INP derivative constituting | + | studies have shown that an INP derivative constituting the N-and |
- | the | + | C-terminal |
- | N-and C-terminal domains can and has been used to display foreign | + | domains can and has been used to display foreign proteins on the |
- | proteins on | + | surface of <i>E. |
- | + | coli. </i>(9).In our project we are intending to harness and make use | |
- | harness | + | of this |
- | and make use of this feature by fusing a specific protein to it. <br> | + | feature by fusing a specific protein to it. <br> |
<i>We have modified this protein to be consistent with BBF RFC-12 | <i>We have modified this protein to be consistent with BBF RFC-12 | ||
- | Standard.<br> | + | Standard. We |
+ | have submitted this part to the part registry.<br> | ||
For more information go to: <a | For more information go to: <a | ||
href="http://partsregistry.org/wiki/index.php/Part:BBa_K265008"><span | href="http://partsregistry.org/wiki/index.php/Part:BBa_K265008"><span | ||
- | style="color: blue;">BBa_K265008</span></a> | + | style="color: blue;">BBa_K265008</span></a> </i><o:p></o:p></span></p> |
- | < | + | <p class="MsoNormal" style="line-height: normal;"><span |
- | style=" | + | style="font-size: 12pt; font-family: "Times New Roman","serif";"> </span><a |
- | + | name="SS"></a><b><span | |
- | style="font-size: 12pt; font-family: "Times New Roman","serif";"> | + | style="font-size: 14pt; font-family: "Times New Roman","serif";">SS</span></b><b><span |
- | < | + | style="font-size: 13pt; font-family: "Times New Roman","serif";">:</span></b><span |
- | <p class="MsoNormal" style="line-height: normal;"><a name="OmpA"></a><b><span | + | style="font-size: 13pt; font-family: "Times New Roman","serif";"> </span><span |
- | style="font-size: 13pt; font-family: "Times New Roman","serif";">OmpA</span></b><span | + | style="font-size: 12pt; font-family: "Times New Roman","serif";">It |
+ | allows protein fused to outer | ||
+ | membrane proteins to become cleaved free. In this experiment this | ||
+ | signal | ||
+ | sequence when placed between INPNC, contains a cleavable site that | ||
+ | allows the | ||
+ | target fusion protein to ‘secrete’ from INPNC. We will do the same with | ||
+ | OmpA. <br> | ||
+ | <i>We have modified this protein to be consistent with BBF RFC-12 | ||
+ | Standard. We | ||
+ | have submitted this part to the part registry.<br> | ||
+ | For more information go to: <a | ||
+ | href="http://partsregistry.org/wiki/index.php/Part:BBa_K265002"><span | ||
+ | style="color: blue;">BBa_K265002</span></a></i><o:p></o:p></span></p> | ||
+ | <p class="MsoNormal" style="line-height: normal;"><a name="INPNCSS"></a><b><span | ||
+ | style="font-size: 13.5pt; font-family: "Times New Roman","serif";">INPNC | ||
+ | + SS: </span></b><span | ||
+ | style="font-size: 12pt; font-family: "Times New Roman","serif";">It | ||
+ | has been suggested that INPNC can | ||
+ | secrete the interest gene better if it is used with the right signal | ||
+ | sequence. Therefore, we are using INPNC and SS toegther in one of our | ||
+ | secretion models.<br> | ||
+ | <i>We have submitted this part to the part registry.<br> | ||
+ | For more information go to: <a | ||
+ | href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K265009"><span | ||
+ | style="color: blue;">BBa_K265009</span></a><o:p></o:p></i></span></p> | ||
+ | <p class="MsoNormal" style="line-height: normal;"><a name="OmpAss"></a><b><span | ||
+ | style="font-size: 13.5pt; font-family: "Times New Roman","serif";">OmpA | ||
+ | + SS: </span></b><span | ||
+ | style="font-size: 12pt; font-family: "Times New Roman","serif";">Since | ||
+ | OmpA is believed to function | ||
+ | similarly to INPNC and INPNC believed to function better when it is | ||
+ | used with | ||
+ | the right signal sequence, we have decided to test and see if OmpA's | ||
+ | ability to | ||
+ | secret increases when it is used by a signal sequence.<br> | ||
+ | <i>We have submitted this part to the part registry.</i><br> | ||
+ | <i>For more information go to: </i><a | ||
+ | href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K265011"><i><span | ||
+ | style="color: blue;">BBa_K265011</span></i></a><o:p></o:p></span></p> | ||
+ | <hr style="width: 100%; height: 2px;"><b><span | ||
+ | style="font-size: 13pt; font-family: "Times New Roman","serif";"><a | ||
+ | name="OmpA"></a>OmpA</span></b><span | ||
style="font-size: 18pt; font-family: "Times New Roman","serif";">:</span><span | style="font-size: 18pt; font-family: "Times New Roman","serif";">:</span><span | ||
style="font-size: 13pt; font-family: "Times New Roman","serif";"> </span><span | style="font-size: 13pt; font-family: "Times New Roman","serif";"> </span><span | ||
Line 157: | Line 218: | ||
system (via fusion with a target | system (via fusion with a target | ||
protein | protein | ||
- | linked with a cleavable signal sequence) <o:p></o:p></span | + | linked with a cleavable signal sequence) <o:p></o:p></span><span |
- | + | style="font-size: 12pt; font-family: "Times New Roman","serif";"><i><br> | |
- | style="font-size: 12pt; font-family: "Times New Roman","serif";">< | + | We |
- | + | have modified this protein to be consistent with BBF RFC-12 Standard.</i></span><i><span | |
- | have modified this protein to be consistent with BBF RFC-12 Standard.</i></span>< | + | style="font-family: "Times New Roman","serif";"><br> |
- | <span style=" | + | </span>Note: |
“It has remained essentially unknown how proteins of E. coli | “It has remained essentially unknown how proteins of E. coli | ||
outer | outer | ||
- | membrane are sorted and incorporated into this membrane” (10)</i> < | + | membrane are sorted and incorporated into this membrane” (10)</i> <span |
- | <i>For more information go to:<a | + | style="font-size: 12pt; font-family: "Times New Roman","serif";"><i><br> |
+ | For more information go to:<a | ||
href="http://partsregistry.org/wiki/index.php/Part:BBa_K103006"><span | href="http://partsregistry.org/wiki/index.php/Part:BBa_K103006"><span | ||
- | style="color: blue;"> BBa_K103006</span></a></i> <o:p></o:p></span | + | style="color: blue;"> BBa_K103006</span></a></i> <o:p></o:p></span> |
<div class="MsoNormal" | <div class="MsoNormal" | ||
style="margin-bottom: 0.0001pt; text-align: center; line-height: normal;" | style="margin-bottom: 0.0001pt; text-align: center; line-height: normal;" | ||
Line 250: | Line 312: | ||
style="color: blue;"> BBa_R0010</span></a><br style=""> | style="color: blue;"> BBa_R0010</span></a><br style=""> | ||
<!--[endif]--></i><o:p></o:p></span></p> | <!--[endif]--></i><o:p></o:p></span></p> | ||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
- | |||
<div class="MsoNormal" | <div class="MsoNormal" | ||
style="margin-bottom: 0.0001pt; text-align: center; line-height: normal;" | style="margin-bottom: 0.0001pt; text-align: center; line-height: normal;" |
Revision as of 21:37, 17 October 2009
Parts related to secretion: Parts related to pH sensor:
Proteins: |
Promoters: |
Others: |
New parts: |
Promoters: |
Proteins: |
New parts:
INPNC:
An
exogenous gene, Ice-nucleation
protein (INP) from Pseudomonas syringae was suggested to be
used for
display of foreign proteins on the surface of E. coli. (7).
Furthermore,
studies have shown that an INP derivative constituting the N-and
C-terminal
domains can and has been used to display foreign proteins on the
surface of E.
coli. (9).In our project we are intending to harness and make use
of this
feature by fusing a specific protein to it.
We have modified this protein to be consistent with BBF RFC-12
Standard. We
have submitted this part to the part registry.
For more information go to: BBa_K265008
SS: It
allows protein fused to outer
membrane proteins to become cleaved free. In this experiment this
signal
sequence when placed between INPNC, contains a cleavable site that
allows the
target fusion protein to ‘secrete’ from INPNC. We will do the same with
OmpA.
We have modified this protein to be consistent with BBF RFC-12
Standard. We
have submitted this part to the part registry.
For more information go to: BBa_K265002
INPNC
+ SS: It
has been suggested that INPNC can
secrete the interest gene better if it is used with the right signal
sequence. Therefore, we are using INPNC and SS toegther in one of our
secretion models.
We have submitted this part to the part registry.
For more information go to: BBa_K265009
OmpA
+ SS: Since
OmpA is believed to function
similarly to INPNC and INPNC believed to function better when it is
used with
the right signal sequence, we have decided to test and see if OmpA's
ability to
secret increases when it is used by a signal sequence.
We have submitted this part to the part registry.
For more information go to: BBa_K265011
OmpA: OmpA is one of the proteins on the outer membrane of E. coli (13),it is used as a displaying fusion protein on the cell surface . This part has already been documented on the parts registry; however, it has not been tested as a compnent of secretion system (via fusion with a target protein linked with a cleavable signal sequence)
We have modified this protein to be consistent with BBF RFC-12 Standard.
Note: “It has remained essentially unknown how proteins of E. coli outer membrane are sorted and incorporated into this membrane” (10)
For more information go to: BBa_K103006
RBS:
Ribosome
Binding site number 32 (BBa_J61132)
from the registry is being used in our secretion system.
For more information go to: BBa_J61132
Terminator: We
are using BBa_B0015, a double terminator, as
our terminator in both our secretion and pH system.
For more information go to: BBa_B0015
GFP (Green
Fluorescent Protein) :
Mutant of GFP known to be very stable (superfolder), which will let
this protein
fold quickly so we can use either a fluorescent reader or UV light to
detect it. Therefore it has been used as a reporter in our secretion
system.
It also serves as a small protein in testing our secretion system.
For more informaiton go to: BBa_K265003
Luciferase: Luciferase
is a firefly protein that also
fluoresces, so it serves as a reporter as well as a testable large
protein.
For more information go to: BBa_1712019
LacI: One
inducible Promoter which was found in the
part registry.
For more information go to: BBa_R0010
6-His
Tag:The
6-Histidine Tag serves as a tag for Western
Blotting if our fluorescent reporters are not expressed as highly as we
would
like.
Note: We are using this tag, just in case if the GFP or Luciferase
does not
work under a plate reader.
ChvI
promoter: Gene
fusion studies confirmed that ChvI gene
was induced by acidic conditions (1). Also, it has been known to
implicate in
virulence (1). This gene is one of the candidates to be use in our
biological
pH sensor as a promoter.
KatA
promoter :This
Chromosomal gene is located on the linear
chromosome (2) and it seems to be induced under an acidic environment
as well
as being involved in the Agrobacterium tumorigenesis
(2).Research has
suggested that ChvG is needed for "responsiveness of gene
expression
to low pH "(2). This gene has become a candidate to complete our pH
sensor
device from this evidence.
AopB
promoter: This
Chromosomal gene located on the circular
chromosome (2) encodes an outer member protein exposed on the bacterial
cell
surface (2). Also, ChvG was shown to be absolutely required for this
gene
expression (2)It seems to get induced under an acidic environment as
well as
being involved in the Agrobacterium tumorigenesis (2).
Therefore,
we have chosen this gene to be one of our candidates to complete our pH
sensor
device.
PhoA
promoter: There
has been a suggestion that ChvI can
activate AP activity by activating transcription of this gene, PhoA
(3).
Therefore, this gene has become one of our candidates to complete our
pH sensor
device.
ImpA
promoter:Gene
fusion studies confirmed that impA genes
was induced by acidic conditions (1), therefore, this is one of our
candidates
to complete our pH sensor device.