Team:Davidson-MissouriW/MeasuringExpression
From 2010.igem.org
(Difference between revisions)
Theshu0012 (Talk | contribs) |
Theshu0012 (Talk | contribs) |
||
Line 8: | Line 8: | ||
<p> Insert text here</p> | <p> Insert text here</p> | ||
- | < | + | <h3> Results from past Davidson and Missouri Western research hinted at the presence of a transcriptional terminator within the TetA gene. Any gene located downstream of the 3’ end of the TetA gene did not appear to be expressed. This gene codes for an effluent pump that physically removes the antibody tetracycline from the cell. |
To test for the presence of this terminator, we created a construct with a fluorescent protein gene downstream of TetA. If TetA really does block transcription, we would expect to see no fluorescence despite induction of the promoter that precedes the construct. We also tested this construct to see if the gene products provided protection from tetracycline compared to a negative control. | To test for the presence of this terminator, we created a construct with a fluorescent protein gene downstream of TetA. If TetA really does block transcription, we would expect to see no fluorescence despite induction of the promoter that precedes the construct. We also tested this construct to see if the gene products provided protection from tetracycline compared to a negative control. | ||
<img src="https://static.igem.org/mediawiki/igem.org/f/f9/Davidson-MissouriWTPS1.png" id="fig1" height=600 width = 800 onclick= "makeBig('fig1')"> | <img src="https://static.igem.org/mediawiki/igem.org/f/f9/Davidson-MissouriWTPS1.png" id="fig1" height=600 width = 800 onclick= "makeBig('fig1')"> | ||
Line 14: | Line 14: | ||
<img src="https://static.igem.org/mediawiki/igem.org/3/3a/Davidson-MissouriWTPS2.png" id="fig2" height=600 width = 800> | <img src="https://static.igem.org/mediawiki/igem.org/3/3a/Davidson-MissouriWTPS2.png" id="fig2" height=600 width = 800> | ||
We performed another battery of tests to compare the pLac+TetA+RFP construct to a construct with the opposite orientation. When compared to cells with RFP followed by TetA, the construct’s negligible fluorescence was even more apparent. Interestingly, the cells with the flipped orientation grew poorly in the presence of tetracycline but had a high fluorescence per cell. One possible explanation for this phenomenon is that the fluorescence is divided by such a low number of cells that small variations are magnified. Alternatively, the presence of tetracycline could be selecting for cells with high expression levels of both gene products. These few cells are producing high levels of TetA effluent pumps in order to survive. Correspondingly, they are producing a lot of RFP. | We performed another battery of tests to compare the pLac+TetA+RFP construct to a construct with the opposite orientation. When compared to cells with RFP followed by TetA, the construct’s negligible fluorescence was even more apparent. Interestingly, the cells with the flipped orientation grew poorly in the presence of tetracycline but had a high fluorescence per cell. One possible explanation for this phenomenon is that the fluorescence is divided by such a low number of cells that small variations are magnified. Alternatively, the presence of tetracycline could be selecting for cells with high expression levels of both gene products. These few cells are producing high levels of TetA effluent pumps in order to survive. Correspondingly, they are producing a lot of RFP. | ||
- | </ | + | </h3> |
</div> | </div> | ||
Revision as of 12:56, 28 July 2010
iGEM Davidson – MWSU 2010: PUT HEADER HERE
Insert text here