Difference between revisions of "Team:MIT/Experiments/Repressors"
| Line 12: | Line 12: | ||
<h2> Purpose</h2> | <h2> Purpose</h2> | ||
| − | <p> | + | <p> Our goal in this experiment was to compare the activity of the repressors BM3R1, TAL14, and TAL21. </p> |
<h2> Set Up </h2> | <h2> Set Up </h2> | ||
| − | <p>We created a plasmid containing each repressor under the control of | + | <p>We created a plasmid containing each repressor under the control of the doxycycline-inducible promoter TRE, which allowed us to modulate the repressor expression by varying the concentration of doxycycline added. We also cloned a repressible promoter controlling the expression of a fluorescent protein. This plasmid allowed us to measure the amount of repression occurring. </p> |
<br> | <br> | ||
<img src="https://static.igem.org/mediawiki/2016/a/ab/MIT_repressor_expt.png" alt="Plasmid Layout"> | <img src="https://static.igem.org/mediawiki/2016/a/ab/MIT_repressor_expt.png" alt="Plasmid Layout"> | ||
<h2> Results </h2> | <h2> Results </h2> | ||
| − | <p>In order for us to analyze the level of repression, we need to see sustained presence of the repressor. Upon increasing the concentration of doxycycline, we were unable to see that sustained presence of the repressors. From the results of testing BM3R1 below, it can be seen that although there is activation early on, the production of repressor falls at 500 nM. This could | + | <p>In order for us to analyze the level of repression, we need to see sustained presence of the repressor. Upon increasing the concentration of doxycycline, we were unable to see that sustained presence of the repressors. From the results of testing BM3R1 below, it can be seen that although there is activation early on, the production of repressor falls at 500 nM. This could possibly be due to errors in doxycycline dilutions at 500 nM and above. |
</p> | </p> | ||
<img src="https://static.igem.org/mediawiki/2016/e/e0/MIT_repressor_results.png" alt="Repressor Experiment Results"> | <img src="https://static.igem.org/mediawiki/2016/e/e0/MIT_repressor_results.png" alt="Repressor Experiment Results"> | ||
Latest revision as of 01:45, 20 October 2016
Testing Repressors
Purpose
Our goal in this experiment was to compare the activity of the repressors BM3R1, TAL14, and TAL21.
Set Up
We created a plasmid containing each repressor under the control of the doxycycline-inducible promoter TRE, which allowed us to modulate the repressor expression by varying the concentration of doxycycline added. We also cloned a repressible promoter controlling the expression of a fluorescent protein. This plasmid allowed us to measure the amount of repression occurring.
Results
In order for us to analyze the level of repression, we need to see sustained presence of the repressor. Upon increasing the concentration of doxycycline, we were unable to see that sustained presence of the repressors. From the results of testing BM3R1 below, it can be seen that although there is activation early on, the production of repressor falls at 500 nM. This could possibly be due to errors in doxycycline dilutions at 500 nM and above.
