Part:BBa_K649105:Experience

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Applications of BBa_K649105

Median fluorescence intensity(MFI) is decreased by LsrR repression.
This work is done by Hiroki Yoshise.

We confirmed that LsrR represses lsrA promoter.For this purpose, we constructed BBa_K649105(PlsrA-gfp-PlsrR-lsrR) and compared fluorescence intensity levels of BBa_K649104(PlsrA-RBS-gfp) and BBa_K649105.As a consequence, Median fluorescence intensity(MFI) of BBa_K649104(PlsrA-RBS-gfp) is 3-fold higher than that of BBa_K649105(PlsrA-gfp-PlsrR-lsrR). This result shows that LsrR represses lsrA promoter and BBa_K649105 works.

[Sample]

pSB6A1 Ptet-GFP RBS1-12(JM2.300)(positive control)

pSB6A1 ⊿P-GFP(JM2.300)(negative control)

pSB3K3 PlsrA-GFP(MG1655)

pSB3K3 PlsrA-GFP-PlsrR-lsrR(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 or 1:100.

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.

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