Part:BBa_K1072000:Experience

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Applications of BBa_K1072000

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[http://2016.igem.org/Team:IISc_Bangalore]Team IISc Bangalore

Fluorescence/OD versus time curve for GFP expression under lux QS

While several quorum sensing parts exist in Parts Registry, relatively few examples exist of a full quorum-sensing sender and receiver in the same plasmid. BBa_K1072000 consists of both a Lux-based sender and a receiver module with GFPmut3 as the reporter. While the part has been shown to express GFPmut3 via microscopy, no growth curves of the same had been reported so far. Therefore, the IISc iGEM team 2016 decided to test the quorum sensing of this part in E coli BL21 (DE3).
Protocol:
Four 25 ml of LB media in conical flasks were inoculated from an overnight primary culture of E. coli BL21 (DE3) transformed with appropriate part (BBa_K1072000 for lux part and BBa_R0040 for negative control) in pSB1C3 (1 % inoculation, 37 degree celcius, 180 RPM). The cultures were grown in an orbit shaker incubator at 37 degree celcius, 180 RPM. 200 uL aliquots were taken from each conical flask every 2 hours and stored at 4 degree celcius. GFPmut3 fluorescence (Ex: 501 nm, Em: 511 nm) and OD600 of the samples were recorded on a plate reader (Tecan M2000 InfinitePro). (For detailed measurement conditions, please refer to the <raw data file>.)
Results
Our experiment confirms the occurrence of a switch due to the QS system in BBa_1072000. This allows us to hypothesize practical applications of this system for cell density dependent processes, such as cell-density dependent protein over-expression for our project.