UC San Diego

An important transposable element (transposon) within the genome of the Escherichia coli K-12 strain BW25113 is IS5, of which there are 11 total copies. This work follows up on past research into IS5 by investigating which one of these 11 copies is the most significant and most impactful on overall IS5 activity and transposition. In addition, it investigates possible environmental and genomic factors influencing the most significant copies, as well as the mechanism of transposition (“cut and paste” or “copy and paste”) used by these copies. We do so by the use of mutation assays, in which strains to be evaluated are plated on minimal media such that the only way for bacterial colonies to grow is for certain IS5 transpositions to occur, making the number of resulting growing colonies a useful indicator of overall IS5 activity. We find that, by far, the most significant copy of IS5 is the copy at the nmpC location, followed by copies at the lnt and wbbL locations, and that a further two copies have mutations that prevent IS5 transposition. We find that the significance of these locations remains invariant with respect to the operon and minimal media used in the mutation assay. We also examine the nmpC copy in more detail and find the reason for its significance is likely due to the presence of a very strong promoter upstream of this gene, and that there is minimal evidence for a bias of IS5 transposases towards acting on this copy. We construct a strain with only the IS5 copy at nmpC and establish a “copy and paste” mechanism for IS5 transposition from nmpC. Lastly, we investigate the possible roles that two genes that overlap the ins5a transposase gene, ins5b and ins5c, play in regulating IS5 transposition, and find that higher levels of their expression have minimal effect.