Difference between revisions of "Part:BBa J64010:Experience"
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<I>Tokyo-Tech iGEM 2011</I> | <I>Tokyo-Tech iGEM 2011</I> | ||
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− | Fluorescence intensity of PlasI(BBa_J64010)-gfp did not change before and after | + | Fluorescence intensity of PlasI(BBa_J64010)-gfp did not change before and after 3OC12-HSL induction. |
Revision as of 02:29, 3 October 2011
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UNIQd02efab4ce2478e0-partinfo-00000000-QINU UNIQd02efab4ce2478e0-partinfo-00000001-QINU
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Tokyo-Tech iGEM 2011 |
Fluorescence intensity of PlasI(BBa_J64010)-gfp did not change before and after 3OC12-HSL induction.
Since Ptrc is a constitutive promoter, lasR is constitutively expressed and, since it is know that this lasR is a working part, the fact that the fluorescence intensity levels do not change before and after induction of BBa_K649000 by 3OC12-HSL indicates that BBa_K649000 is not regulated by 3OC12-HSL.
PlasI(BBa_J64010)-rbs-gfp-TT / Ptrc-rbs-lasR-TT
①Overnight culture of grown at 37 in LB medium containing carbenicillin and kanamycin were diluted 1:1000 in the medium, and then they were incubated at 37 °C as fresh cultures. ②After their OD600 reached 0.2, we added 3 µL of 500 µM 3O-C12-HSL (3OC12-HSL+) or 3µL of DMSO (3OC12-HSL-) into the fresh cultures. ③After 3-hour incubation at 37 °C (OD reached approximately 1.50.), 0.25 mL of each culture was harvested by centrifugalization and suspended by adding 1 mL of PBS (phosphate-buffered saline). ④We dispensed 500 µL of each suspension into a disposable tube through a cell strainer, and its fluorescence intensity was measured with a flow cytometer of Becton, Dickinson and Company. |