Part:BBa_K639003:Experience

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Imperial iGEM 2013

Stress biosensor characterisation (BBa_K639003)

Originally we intended on using BBa_K639003 to detect whether our cells were stressed when placed in various toxic byproducts. However, as the data below shows, this biobrick is very leaky. As such, as an alternative we utilised the stress sensor as a marker for our chassis E. coli (MG1655) for an array of qualitative and quantitative waste growth and toxicity assays.

mCherry production is induced by the stress pathway and detection of ppGpps. Induction with IPTG bypassess this mechanism through an inhibition of LacI, resulting in mCherry expression.

Stress sensor growth assayE. coli (MG1655) transformed with stress biosensor (BBa_K639003) and grown at 37oC, with shaking for 6 hours. Error bars represent S.E.M. n=4	Stress sensor IPTG induced fluorescenceE. coli (MG1655) transformed with stress biosensor (BBa_K639003) and grown at 37oC, with shaking for 6 hours. Error bars represent S.E.M. n=4
BBa_K639003 transformed into E coli. strain MG1655. Pink colonies are visible, which relate to 'leaky' mCherry production	BBa_K639003 transformed into E coli. strain MG1655. Cells were grown at 37oC in 4ml LB with 0, 1 or 2mM IPTG. At 6 hours post IPTG induction, cells were spun down and imaged.