Part:BBa_K1149051:Experience

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UNIQ92dead5d77669cef-partinfo-00000000-QINU UNIQ92dead5d77669cef-partinfo-00000001-QINU The Stanford-Brown 2015 team wanted to build off of the Imperial College team's work by adding the gene for a type II pantothenate kinase (panK) to this part to make a composite part (BBa_K1692021). To test whether the panK gene caused more plastic to be produced, we used this BioBrick as a comparison. Our final results did show that the addition of the panK gene did cause an increase in plastic production, as shown by the figure below.

This plot shows the increase of the percent of dry cell mass that is P(3HB) and the increase in relative fluorescence. The measurements of our construct are in general higher by both of these measurements, which suggests that our construct works successfully.

Notably, we used the strain NEB5-alpha instead of the same strain as the Imperial 2013 team used (MG1655). NEB5-alpha is not known to be a high accumulator of P(3HB), but is is a common strain so we wanted to use it to test our construct. Even though our construct produced, on average, a 23% increase in the amount of plastic in vivo as a percentage of dry cell weight over the Imperial 2013 construct in our lab, we still produced less plastic than the Imperial 2013 did. As opposed to their value of 58.9% plastic, we were only able to achieve 25-34% yield. We suspect that is due to the strain difference, since the media difference should have worked in our favor (we used TB, which the Tokyo Tech iGEM 2012 team had shown causes more plastic to be produced). However, supplementing the media with glucose, as opposed to as with glycerol, could have also caused the difference in accumulation.