Part:BBa_K649105:Experience

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Applications of BBa_K649105

Fluorescence intensity is decreased by LsrR repression.
This work is done by Hiroki Yoshise.

We confirmed that LsrR represses lsrA promoter. We measured LsrR repression activity by using a plasmid (BBa_K649105) which have a lsrR gene downstream of lsrA promoter-gfp.By the LsrR repression, the fluorescence intensity decreased 3-fold. This result shows that LsrR successfully repressed lsrA promoter. The working part we created allow the use of AI-2 as a signaling molecule, which is a very powerful tool to build complex Synthetic Biology systems.

[Sample]

Ptet-gfp on pSB6A1(JM2.300)(positive control)

Promoterless-gfp on pSB6A1(JM2.300)(negative control)

PlsrA-gfp on pSB3K3(MG1655)

PlsrA-gfp-PlsrR-lsrR on pSB3K3(MG1655)

[Method]

1. Overnight cultures of reporter strains grown at 37 °C in LB medium containing appropriate antibiotics were diluted 1:100 into 3 ml of LB medium and were incubated at 37 °C as fresh cultures.

2. After their OD590 reached 0.15, the fresh cultures were diluted 1:10.

3. After 4-hour incubation at 37 °C, 1 ml of each culture was moved to 1.6ml tube and its fluorescence intensity was measured with a flow cytometer.

After four hours from OD590 reaching 0.15, we measured OD.
This work is done by Hiroki Yoshise.

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