Difference between revisions of "Part:BBa K5097002:Experience"

Latest revision as of 06:09, 2 October 2024

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Applications of BBa_K5097002

Experience

We tested the expression of BFP under control of PREm13. To do this, we generated a reporter construct (BBa_K5097005). We used this to transform E. coli DH5alpha cells, and then grew the cells in LB supplemented with buffering agents to achieve the desired pH. In parallel, we measured the OD595 of the culture and its fluorescence (Ex. 374 nm, Em. 446 nm) We then normalized the absorbance to cell density and calculated the change in this normalized value over the course of the experiment.

Figure 3: Expression of BFP under the control of PREm13 in media with different pH. The pH indicated was created by adding a buffering agent to LB to adjust the pH. UB, unbuffered LB.

From these results we concluded that the PREm13 riboswitch does display pH sensitive induction of BFP, with expression maximally at pH 9, consistent with the activity reported of the Alx riboswitch, which is induced maximally at pH 8.5.

User Reviews

UNIQ239bcc836b1c3e32-partinfo-00000000-QINU UNIQ239bcc836b1c3e32-partinfo-00000001-QINU

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 ===Experience===
-Since the structure of the riboswitch was maintained, despite the presence of three deletions, we tested its ability to regulate translation of BFP in response to pH changes. To do this, we generated a reporter construct BBa_K5097005. We used this to transform E. coli DH5alpha cells, and then grew the cells in LB supplemented with buffering agents to achieve the desired pH. In parallel, we measured the OD595 of the culture and its fluorescence (Ex. 374 nm, Em. 446 nm).  We then normalized the absorbance to cell density and calculated the change in this normalized value over the course of the experiment.
+We tested the expression of BFP under control of PREm13.  To do this, we generated a reporter construct (BBa_K5097005). We used this to transform E. coli DH5alpha cells, and then grew the cells in LB supplemented with buffering agents to achieve the desired pH. In parallel, we measured the OD595 of the culture and its fluorescence (Ex. 374 nm, Em. 446 nm)  We then normalized the absorbance to cell density and calculated the change in this normalized value over the course of the experiment.
 https://static.igem.wiki/teams/5097/parts/team-oneonta-2024-prem13-growthstudy.jpg
 ::Figure 3: Expression of BFP under the control of PREm13 in media with different pH.  The pH indicated was created by adding a buffering agent to LB to adjust the pH. UB, unbuffered LB.
-From these results we concluded that the PREm13 riboswitch does display pH sensitive induction of BFP, with expression maximally at pH 9.
+From these results we concluded that the PREm13 riboswitch does display pH sensitive induction of BFP, with expression maximally at pH 9, consistent with the activity reported of the Alx riboswitch, which is induced maximally at pH 8.5.