Difference between revisions of "Team:Uppsala/Phage Display"
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<h1>Experiment</h1> | <h1>Experiment</h1> | ||
<p> | <p> | ||
| − | There are no readily available protocols for whole-organism exterior screenings using phage display, to read more closely what how we adapted this procedure to fit our needs read more here | + | There are no readily available protocols for whole-organism exterior screenings using phage display, to read more closely what how we adapted this procedure to fit our needs read more here. |
<br> | <br> | ||
<br> | <br> | ||
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<br> | <br> | ||
<br> | <br> | ||
| − | Read more about our modified protocol here | + | Read more about our modified protocol here. |
</p> | </p> | ||
| Line 324: | Line 324: | ||
<h1>Results</h1> | <h1>Results</h1> | ||
<p> | <p> | ||
| − | + | Our three parallell pannings and subsequent analysis led to the following peptides being characterized: | |
| + | </p> | ||
| + | <div> | ||
<br> | <br> | ||
| − | < | + | <p><strong>Table 1:</strong> Final peptides and their characteristics. *TUP = Target-Unrelated Peptide. |
| − | + | ||
</p> | </p> | ||
| + | <table class="wikitable"> | ||
| + | <tbody> | ||
| + | <tr> | ||
| + | <th> | ||
| + | </th> | ||
| + | <th> Sample | ||
| + | </th> | ||
| + | <th> Aligned Sequence | ||
| + | </th> | ||
| + | <th> Propagation Advantage | ||
| + | </th> | ||
| + | <th>Predicted Polysterene Binder | ||
| + | </th> | ||
| + | <th> Known TUP*-Motif | ||
| + | </th> | ||
| + | </tr> | ||
| + | <tr> | ||
| + | <th> Series 1: | ||
| + | </th> | ||
| + | <th> EF01122224: TPIFLPTPAQEH | ||
| + | </th> | ||
| + | <th> TP<span class="blue-letter">I</span>FL<span class="red-letter">P</span>TPA<span class="blue-letter">Q</span>EH--- | ||
| + | </th> | ||
| + | <th>Yes | ||
| + | </th> | ||
| + | <th>No | ||
| + | </th> | ||
| + | <th>No | ||
| + | </th> | ||
| + | </tr> | ||
| + | |||
| + | |||
| + | <tr> | ||
| + | <th> Series 3: | ||
| + | </th> | ||
| + | <th> EF01122218: FSPTQANTIHRW | ||
| + | </th> | ||
| + | <th> ---FS<span class="red-letter">P</span>TQA<span class="blue-letter">N</span>T<span class="blue-letter">IH</span>RW | ||
| + | </th> | ||
| + | <th>No | ||
| + | </th> | ||
| + | <th>No | ||
| + | </th> | ||
| + | <th>No | ||
| + | </th> | ||
| + | </tr> | ||
| + | |||
| + | <tr> | ||
| + | <th> | ||
| + | </th> | ||
| + | <th> EF01122220: VGGTVQSESHRR | ||
| + | </th> | ||
| + | <th>--<span class="blue-letter">V</span>GGTVQSESH<span class="blue-letter">R</span>R- | ||
| + | </th> | ||
| + | <th>No | ||
| + | </th> | ||
| + | <th>No | ||
| + | </th> | ||
| + | <th>No | ||
| + | </th> | ||
| + | </tr> | ||
| + | |||
| + | <tr> | ||
| + | <th> | ||
| + | </th> | ||
| + | <th> EF01122222: SMGRTDYVQQLR | ||
| + | </th> | ||
| + | <th>-S<span class="blue-letter">M</span>GRTDYV<span class="blue-letter">Q</span>Q<span class="blue-letter">LR</span>-- | ||
| + | </th> | ||
| + | <th>No | ||
| + | </th> | ||
| + | <th>No | ||
| + | </th> | ||
| + | <th>No | ||
| + | </th> | ||
| + | </tr> | ||
| + | |||
| + | <tr> | ||
| + | <th> | ||
| + | </th> | ||
| + | <th> EF01122217: RVQPAHFNVMGQ | ||
| + | </th> | ||
| + | <th>--RVQ<span class="red-letter">P</span>AHF<span class="blue-letter">N</span>V<span class="blue-letter">M</span>GQ | ||
| + | </th> | ||
| + | <th>No | ||
| + | </th> | ||
| + | <th>Yes | ||
| + | </th> | ||
| + | <th>No | ||
| + | </th> | ||
| + | </tr> | ||
| + | |||
| + | </table> | ||
| + | </div> | ||
</div> | </div> | ||
Revision as of 23:16, 15 October 2018
Design
We used a Phage Display screening to create a biosensor that aggregated around the target molecule with the hypothesis that this would create a larger response.
Phage display is a method that is usually used to find a protein that binds to another protein. However using this technique on a whole eukaryotic organism is very uncommon, and therefore this project represents breaking new ground, albeit with the inevitable cost of hammering out the problems that come with such an undertaking.
In order to make our method work, we had to immobilize our worms and be able to extract the bound phages without also extracting the worms! Since our worms are far larger than a phage, we used an eppendorf filter insert that had pores too small for strongyles to pass through and allowed our phages to bind to these worms.
Using this process, we found several peptide sequences with potential high affinity to our worms. These sequences, after further validation, can in future be used to synthesize E. coli that will bind to our worms!
To read more about the general design of a Phage Display Screening and why we chose to apply this method, click here:
Experiment
There are no readily available protocols for whole-organism exterior screenings using phage display, to read more closely what how we adapted this procedure to fit our needs read more here.
Protocol
For this experiment we used a modified version of the protocol supplied with the kit from New England Biolabs. You can find the original protocol here.
Read more about our modified protocol here.
Results
Our three parallell pannings and subsequent analysis led to the following peptides being characterized:
Table 1: Final peptides and their characteristics. *TUP = Target-Unrelated Peptide.
| Sample | Aligned Sequence | Propagation Advantage | Predicted Polysterene Binder | Known TUP*-Motif | |
|---|---|---|---|---|---|
| Series 1: | EF01122224: TPIFLPTPAQEH | TPIFLPTPAQEH--- | Yes | No | No |
| Series 3: | EF01122218: FSPTQANTIHRW | ---FSPTQANTIHRW | No | No | No |
| EF01122220: VGGTVQSESHRR | --VGGTVQSESHRR- | No | No | No | |
| EF01122222: SMGRTDYVQQLR | -SMGRTDYVQQLR-- | No | No | No | |
| EF01122217: RVQPAHFNVMGQ | --RVQPAHFNVMGQ | No | Yes | No |
Notebook
Here you can find our lab notes, the notes and scribbles we took during the course of our experiments: