Team:William and Mary/Improve

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Improved Parts

For our improved part this year, we improved BBa_K2333413 by switching out original promoter, ribosome binding site and terminator for Mycobacterium compatible parts. BBa_K2333413 is a constitutive mScarlet-I expressing circuit, with BBa_J23100 as its promoter. BBa_K3059424 also expresses mScarlet-I, which is driven by a relatively strong Mycobacterium promoter S16 (BBa_K2333410 ). We achieved similar fluorescence intensity level by transforming BBa_K3059424 into Mycobacterium smegmatis; Normalized plate reader data is plotted in the following graph:

Fluorescence intensity normalized with respect to OD600 plotted on a log scale. Each dot represents a distinct biological replicate (colonies). Negative control measures untransformed, non-fluorescent M.smeg mc2155.

We can see that BBa_K3059424 showed slightly lower fluorescence intensity measurements than BBa_K2333413 , but still highly notable. This circuit could also serve as a framework for engineering mycobacterium using plasmids, since mScarlet-I could easily be switched out to any gene of interest. Please see BBa_K3059424 for more information.

Header picture: Mycobacterium smegmatis visualized under scanning electron microscopy. Source: https://en.wikipedia.org/wiki/Mycobacterium_smegmatis#/media/File:Mycobacterium_smegmatis.tif