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+ | <div class="row"> | ||
+ | |||
+ | <div class="col-sm-7"> | ||
<center><div style='font-size: 22px;padding-bottom: 15px;'> Cloning-Ready Protein Degradation Tags </div></center> | <center><div style='font-size: 22px;padding-bottom: 15px;'> Cloning-Ready Protein Degradation Tags </div></center> | ||
− | <div style='padding-left | + | <div style='padding-left: 10%;text-indent: 30px;'>We utilized a suite of 6 protein degradation tags (pdts) originally designed by [Collins et al 2014]. Each tag confers a distinct level of protein degradation; tags are lettered (A-F) in order of protease affinity, from strongest (highest level of degradation) to weakest. These parts have been made BioBrick compatible, codon-optimized for E. coli, and placed in between UNS sites, so that teams may easily insert them after their gene of choice using Gibson Assembly. They are also flanked by BsaI cut sites, for ease of use with Golden Gate Assembly. This allows future teams to efficiently adapt the speed-control system to their own parts and circuits, regardless of their preferred assembly method.</div> |
<div> | <div> | ||
<img src =""width="200px;"/> | <img src =""width="200px;"/> | ||
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− | + | <div class="col-sm-4"> | |
<center> | <center> | ||
− | <table style=''> | + | <table style='float: right;'> |
<tr> | <tr> | ||
− | <th style='background-color: #BEB9C7;column-width: | + | <th style='background-color: #BEB9C7;column-width: 70px;'><center>Part</center></th> |
− | <th style='background-color: #BEB9C7;column-width: | + | <th style='background-color: #BEB9C7;column-width: 400px;'><center>Description</center></th> |
</tr> | </tr> | ||
<tr> | <tr> | ||
− | <td style='background-color: #9892A1;column-width: | + | <td style='background-color: #9892A1;column-width: 70px; font-size: 15px;'><a href="http://parts.igem.org/Part:BBa_K2333401"> K2333401 </a></td> |
− | <td style='background-color: #ECE7F2;column-width: | + | <td style='background-color: #ECE7F2;column-width: 400px; font-size: 15px;'>Cloning Ready pdt #3A with Double Terminator</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
− | <td style='background-color: #9892A1;column-width: | + | <td style='background-color: #9892A1;column-width: 70px; font-size: 15px;'><a href="http://parts.igem.org/Part:BBa_K2333402"> K2333402 </a></td> |
− | <td style='background-color: #DED9E5;column-width: | + | <td style='background-color: #DED9E5;column-width: 400px; font-size: 15px;'>Cloning Ready pdt #3B with Double Terminator</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
− | <td style='background-color: #9892A1;column-width: | + | <td style='background-color: #9892A1;column-width: 70px; font-size: 15px;'><a href="http://parts.igem.org/Part:BBa_K2333403"> K2333403 </a></td> |
− | <td style='background-color: #ECE7F2;column-width: | + | <td style='background-color: #ECE7F2;column-width: 400px; font-size: 15px;'>Cloning Ready pdt #3C with Double Terminator</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
− | <td style='background-color: #9892A1;column-width: | + | <td style='background-color: #9892A1;column-width: 70px; font-size: 15px;'><a href="http://parts.igem.org/Part:BBa_K2333404"> K2333404 </a></td> |
− | <td style='background-color: #DED9E5;column-width: | + | <td style='background-color: #DED9E5;column-width: 400px; font-size: 15px;'>Cloning Ready pdt #3D with Double Terminator</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
− | <td style='background-color: #9892A1;column-width: | + | <td style='background-color: #9892A1;column-width: 70px; font-size: 15px;'><a href="http://parts.igem.org/Part:BBa_K2333405"> K2333405 </a></td> |
− | <td style='background-color: #ECE7F2;column-width: | + | <td style='background-color: #ECE7F2;column-width: 400px; font-size: 15px;'>Cloning Ready pdt #3D with Double Terminator</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
− | <td style='background-color: #9892A1;column-width: | + | <td style='background-color: #9892A1;column-width: 70px; font-size: 15px;'><a href="http://parts.igem.org/Part:BBa_K2333406"> K2333406 </a></td> |
− | <td style='background-color: #DED9E5;column-width: | + | <td style='background-color: #DED9E5;column-width: 400px; font-size: 15px;'>Cloning Ready pdt #3F with Double Terminator</td> |
</tr> | </tr> | ||
</table> | </table> | ||
</center> | </center> | ||
+ | </div> | ||
+ | <div class="col-sm-1"></div> | ||
− | + | </div> <!----out of row--> | |
− | + | ||
− | <!----out of row--> | + | |
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+ | <div class="row"> | ||
− | + | <div class="col-sm-7"> | |
<center><div style='font-size: 22px;padding-bottom: 15px;'> Constitutive Tagged Reporters </div></center> | <center><div style='font-size: 22px;padding-bottom: 15px;'> Constitutive Tagged Reporters </div></center> | ||
− | <div style='padding-left | + | <div style='padding-left: 10%;text-indent: 30px;'>Our next collection of parts consist of a protein degradation tagged mScarlet reporter under the control of the strong constitutive promoter J23100. These parts, in combination with inducible mf-Lon protease constructs (listed below), allowed us to characterize the degradation properties of each protein degradation tag on a plasmid-based system. We successfully demonstrated distinct levels of protein degradation by each of the 6 pdt’s; see our full characterization here [HYPERLINK TO RESULTS PAGE]. We also included a tagless control construct (J23100 mScarlet with no pdt) as a comparison. In order to demonstrate that our protein degradation tags operated similarly regardless of the tagged protein, we also built and characterized analogous constructs with an sfGFP reporter; these are listed below in the “All Parts” table. mScarlet and sfGFP reporters have been codon-optomized for E. coli and feature a double stop codon for ….(enhanced efficiency or something idk). </div> |
<div> | <div> | ||
<img src =""width="200px;"/> | <img src =""width="200px;"/> | ||
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− | + | <div class="col-sm-4"> | |
<center> | <center> | ||
<table style=''> | <table style=''> | ||
<tr> | <tr> | ||
− | <th style='background-color: #BEB9C7;column-width: | + | <th style='background-color: #BEB9C7;column-width: 50px;'><center>Part</center></th> |
− | <th style='background-color: #BEB9C7;column-width: | + | <th style='background-color: #BEB9C7;column-width: 250px;'><center>Description</center></th> |
</tr> | </tr> | ||
<tr> | <tr> | ||
− | <td style='background-color: #9892A1;column-width: | + | <td style='background-color: #9892A1;column-width: 50px; font-size: 15px;'><a href="http://parts.igem.org/Part:BBa_K2333413"> K2333413 </a></td> |
− | <td style='background-color: #ECE7F2;column-width: | + | <td style='background-color: #ECE7F2;column-width: 250px; font-size: 15px;'>J23100 mScarlet-I</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
− | <td style='background-color: #9892A1;column-width: | + | <td style='background-color: #9892A1;column-width: 50px; font-size: 15px;'><a href="http://parts.igem.org/Part:BBa_K2333414"> K2333414 </a></td> |
− | <td style='background-color: #DED9E5;column-width: | + | <td style='background-color: #DED9E5;column-width: 250px; font-size: 15px;'>J23100 mScarlet-I pdt #3A</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
− | <td style='background-color: #9892A1;column-width: | + | <td style='background-color: #9892A1;column-width: 50px; font-size: 15px;'><a href="http://parts.igem.org/Part:BBa_K2333415"> K2333415 </a></td> |
− | <td style='background-color: #ECE7F2;column-width: | + | <td style='background-color: #ECE7F2;column-width: 250px; font-size: 15px;'>J23100 mScarlet-I pdt #3B</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
− | <td style='background-color: #9892A1;column-width: | + | <td style='background-color: #9892A1;column-width: 50px; font-size: 15px;'><a href="http://parts.igem.org/Part:BBa_K2333416"> K2333416 </a></td> |
− | <td style='background-color: #DED9E5;column-width: | + | <td style='background-color: #DED9E5;column-width: 250px; font-size: 15px;'>J23100 mScarlet-I pdt #3C</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
− | <td style='background-color: #9892A1;column-width: | + | <td style='background-color: #9892A1;column-width: 50px; font-size: 15px;'><a href="http://parts.igem.org/Part:BBa_K2333417"> K2333417 </a></td> |
− | <td style='background-color: #ECE7F2;column-width: | + | <td style='background-color: #ECE7F2;column-width: 250px; font-size: 15px;'>J23100 mScarlet-I pdt #3D</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
− | <td style='background-color: #9892A1;column-width: | + | <td style='background-color: #9892A1;column-width: 50px; font-size: 15px;'><a href="http://parts.igem.org/Part:BBa_K2333418"> K2333418 </a></td> |
− | <td style='background-color: #DED9E5;column-width: | + | <td style='background-color: #DED9E5;column-width: 250px; font-size: 15px;'>J23100 mScarlet-I pdt #3E</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
− | <td style='background-color: #9892A1;column-width: | + | <td style='background-color: #9892A1;column-width: 50px; font-size: 15px;'><a href="http://parts.igem.org/Part:BBa_K2333419"> K2333419 </a></td> |
− | <td style='background-color: #DED9E5;column-width: | + | <td style='background-color: #DED9E5;column-width: 250px; font-size: 15px;'>J23100 mScarlet-I pdt #3F</td> |
</tr> | </tr> | ||
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</div> | </div> | ||
+ | <div class="col-sm-1"></div> | ||
− | + | </div> <!----out of row--> | |
− | <!----out of row--> | + | |
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<div class="col-sm-7"> | <div class="col-sm-7"> | ||
− | <center><div style='font-size: 22px;padding-bottom: 15px;'> | + | <center><div style='font-size: 22px;padding-bottom: 15px;'> UNS PDT Parts </div></center> |
− | <div style='padding-left: 10% | + | <div style='padding-left: 10%;'>test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test </div> |
<div> | <div> | ||
<img src =""width="200px;"/> | <img src =""width="200px;"/> | ||
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<div class="col-sm-7"> | <div class="col-sm-7"> | ||
− | <center><div style='font-size: 22px;padding-bottom: 15px;'> | + | <center><div style='font-size: 22px;padding-bottom: 15px;'> UNS PDT Parts </div></center> |
− | <div style='padding-left: 10% | + | <div style='padding-left: 10%;'>test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test </div> |
<div> | <div> | ||
<img src =""width="200px;"/> | <img src =""width="200px;"/> | ||
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<div class="col-sm-7"> | <div class="col-sm-7"> | ||
− | <center><div style='font-size: 22px;padding-bottom: 15px;'> | + | <center><div style='font-size: 22px;padding-bottom: 15px;'> UNS PDT Parts </div></center> |
− | <div style='padding-left: 10% | + | <div style='padding-left: 10%;'>test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test </div> |
− | </div> | + | |
<div> | <div> | ||
<img src =""width="200px;"/> | <img src =""width="200px;"/> |
Revision as of 18:59, 29 October 2017
Submitted Parts to the iGEM Registry
This year we are proud to submit a series of parts allowing for the modular control of gene expression speed, using the mf-Lon/pdt protein degradation system. These parts will enable users to predictably control the temporal dynamical behavior of their gene of choice, incorporating the speed-control system seamlessly into their circuit with our easy-to-clone parts. We include a number of variations in reporter protein and inducible promoter/respective inducer molecule, so that teams can tailor to the requirements of their unique circuit of interest. Additionally, every composite part submitted is flanked by the Unique Nucleotide Sequences (UNS’s) introduced to iGEM by W&M’s 2016 team to improve ease and reliability of cloning.
Cloning-Ready Protein Degradation Tags
We utilized a suite of 6 protein degradation tags (pdts) originally designed by [Collins et al 2014]. Each tag confers a distinct level of protein degradation; tags are lettered (A-F) in order of protease affinity, from strongest (highest level of degradation) to weakest. These parts have been made BioBrick compatible, codon-optimized for E. coli, and placed in between UNS sites, so that teams may easily insert them after their gene of choice using Gibson Assembly. They are also flanked by BsaI cut sites, for ease of use with Golden Gate Assembly. This allows future teams to efficiently adapt the speed-control system to their own parts and circuits, regardless of their preferred assembly method.
K2333401 | Cloning Ready pdt #3A with Double Terminator |
K2333402 | Cloning Ready pdt #3B with Double Terminator |
K2333403 | Cloning Ready pdt #3C with Double Terminator |
K2333404 | Cloning Ready pdt #3D with Double Terminator |
K2333405 | Cloning Ready pdt #3D with Double Terminator |
K2333406 | Cloning Ready pdt #3F with Double Terminator |
Constitutive Tagged Reporters
Our next collection of parts consist of a protein degradation tagged mScarlet reporter under the control of the strong constitutive promoter J23100. These parts, in combination with inducible mf-Lon protease constructs (listed below), allowed us to characterize the degradation properties of each protein degradation tag on a plasmid-based system. We successfully demonstrated distinct levels of protein degradation by each of the 6 pdt’s; see our full characterization here [HYPERLINK TO RESULTS PAGE]. We also included a tagless control construct (J23100 mScarlet with no pdt) as a comparison. In order to demonstrate that our protein degradation tags operated similarly regardless of the tagged protein, we also built and characterized analogous constructs with an sfGFP reporter; these are listed below in the “All Parts” table. mScarlet and sfGFP reporters have been codon-optomized for E. coli and feature a double stop codon for ….(enhanced efficiency or something idk).
UNS PDT Parts
test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test
UNS PDT Parts
test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test
UNS PDT Parts
test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test test
All Submitted Parts
K2333401 | Cloning Ready pdt #3A with Double Terminator |
K2333402 | Cloning Ready pdt #3B with Double Terminator |
K2333403 | Cloning Ready pdt #3C with Double Terminator |
K2333404 | Cloning Ready pdt #3D with Double Terminator |
K2333405 | Cloning Ready pdt #3D with Double Terminator |
K2333406 | Cloning Ready pdt #3F with Double Terminator |
K2333407 | UNS J23100 sfGFP pdt #3A |
K2333408 | UNS J23100 sfGFP pdt #3B |
K2333409 | UNS J23100 sfGFP pdt #3C |
K2333410 | UNS J23100 sfGFP pdt #3D |
K2333411 | UNS J23100 sfGFP pdt #3E |
K2333412 | UNS J23100 sfGFP pdt #3F |
K2333413 | J23100 mScarlet-I |
K2333414 | J23100 mScarlet-I pdt #3A |
K2333415 | J23100 mScarlet-I pdt #3B |
K2333416 | J23100 mScarlet-I pdt #3C |
K2333417 | J23100 mScarlet-I pdt #3D |
K2333418 | J23100 mScarlet-I pdt #3E |
K2333419 | J23100 mScarlet-I pdt #3F |
K2333420 | UNS pTet sfGFP |
K2333421 | UNS pTet sfGFP pdt #3A |
K2333422 | UNS pTet sfGFP pdt #3B |
K2333423 | UNS pTet sfGFP pdt #3C |
K2333424 | UNS pTet sfGFP pdt #3D |
K2333425 | UNS pTet sfGFP pdt #3E |
K2333426 | UNS pTet sfGFP pdt #3F |
K2333427 | UNS pTet mScarlet-I |
K2333428 | UNS pTet mScarlet-I pdt #3A |
K2333429 | UNS pTet mScarlet-I pdt #3B |
K2333430 | UNS pTet mScarlet-I pdt #3C |
K2333431 | UNS pTet mScarlet-I pdt #3D |
K2333432 | UNS pTet mScarlet-I pdt #3E |
K2333433 | UNS pTet mScarlet-I pdt #3F |
K2333434 | pLac0-1 mf-Lon |
K2333435 | pBad mf-Lon |
K2333437 | Copper Sensor pdt #3A |
K2333438 | Copper Sensor pdt #3B |
K2333439 | Copper Sensor pdt #3C |
K2333440 | Copper Sensor pdt #3D |
K2333441 | Copper Sensor pdt #3E |
K2333442 | Copper Sensor pdt #3F |