Part:BBa_K769020:Experience

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Applications of BBa_K769020

Evaluation

Transformed E coli. TOP10 was precultured into LB medium and incubated at 37℃. After incubating for 12 hour, precultured solution was diluted with LB medium to the point that OD660 became 0.6, then applied into 96well microtiter plate. After adding arabinose(0.25% f.c.), the plate was shaken at 20 ℃ to express the lux proteins. The spectrum of the light and intensity was measured by plate reader once in an hour.

Result

We could observe the shift of wavelength from 488 nm to 478 nm (Figure.a). The changing color can clearly recognize by the photograph(Figure.e). And we also achieved to enhance the light intensity. Maximal absorption of Lux+LumP is about 3-fold, comparing to that of lux(cloned). Furthermore, we demonstrated that the lumP part(Edinburgh 2009, BBa_K216007) could work with the Lux operon from Photobacterium phosphoreum kishitanii strain.


The flask on the very right, the brightest strain, is the strain that is expressing both the lux operon and lumP.

User Reviews

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BBa_K769020 iGEM19_Chalmers-Gothenburg

We followed the procedure as given on this page and did the entire experiment over from the start one time. Following the instructions included using the same strain of E.coli, Top10, the same temperature, 20°C and the same concentration of the inducer arabinose, 0.2%. Still we observed no light emission at any point. A sequencing was performed where a missing base pair in the promoter, 44 base pairs before the actual operon, was found. There was also an indication that a base pair in the operon, placed 2337 base pairs into the operon, was missing. This would result in a frameshift. Either of these could potentially be the reason the biobrick did not work as described.

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