Team:Alberta/Project

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Team Alberta

Project Title

Overall Project

A microbial color wheel
This year, the University of Alberta team is developing a biological sensor circuit with three pigment colours to generate a multiple coloured output. We developed this idea because traditional biological reporters have been limited to a small set of fluorescent proteins and colour genes, which produce only an all-or-none output. Furthermore, having only a single channel of output limits the application of other sensors in a single biological device. Therefore, it is clear that the traditional sensor system requires easy-to-use bioreporters that are capable of responding to chemical gradients and mixing independent output channels. To construct the biosensor circuit, we used existing genetic parts pioneered by the 2009 and 2010 iGEM teams, which will result in the circuit being capable of responding to chemical gradients and producing a multi-coloured output in the form of a colour wheel. This biosensor will become the new age of easy–to-read reporters that are incorporated into new versions of the Genomikon kit, which directly impact both research and education usage.

Project Details

Part 2 The Experiments

Part 3

Results

Stage One

June.20.2012

We finished the first part of our biological circuit construction (promoter switching) and discovered that the red fluorescent protein (rfp) gene was only expressed from the two strongest promoters, 4 (second strongest) and 5 (strongest), out of nine different promoters tested. The blue pigment protein (bpp) gene was only expressed from promoter 2, which is the second weakest of the tested promoters..Expression of the green fluorescent protein (gfp) gene was not detected using any promoter, unexpectedly. These results indicate that the recruitment of the RNA polymerase to initiate transcription and expression of these pigment genes can be accomplished using promoter 2 or higher. However, the strength of the promoter does affect visible color development in E. coli colonies. A larger amount of RFP is required as expression was found to only be driven by the strongest promoter while BPP was produced at sufficient levels using a weaker promoter. This may also indicate that BPP is toxic at higher levels driven from stronger promoters. The lack of GFP production may be a result of cloning errors or secondary structure of mRNA blocking RBS from ribosome binding. Therefore, We have decided to construct plasmids to test the different strengths of RBS and investigate the toxicity of colour proteins. We have also begun cloning transcriptional repressors in order to modulate level of pigment gene expression.

Stage Two