Title

Insertion element activity in bacterial genomes

Date of Completion

January 2010

Keywords

Biology, Genetics|Biology, Microbiology|Biology, Bioinformatics

Degree

Ph.D.

Abstract

The ubiquity of Mobile Genetic Elements (MGE) suggests that their continued presence in all domains of life results from a successful evolutionary strategy. The influences of these elements on the genome structure and population dynamics of their host organisms have been an increasing topic of interest in this new age of genome sequencing. I present several bioinformatic and molecular approaches that may serve as guidelines for investigating the influence of a simple type of MGE, the Insertion Sequence (IS), on bacterial genome projects currently in the pipeline. These techniques identify putative and fragmentary IS elements and identify potential genome insertion hotspots. In the case of the root nodule actinobacterium, Frankia sp., ORFs related to IS elements comprise nearly 3% of the genome coding sequences in two of three sequenced strains. Remnant transposase ORFs, detected using a PSI-TBLASTN approach, increased the numbers of predicted IS elements in the HFPCcI3 (CcI3) and EAN1pec (EAN) genomes by 36% and 39% respectively. IS element comparisons using a sliding window analysis revealed clear instances of clustering near regions of predicted gene deletions in strain CcI3 and near regions of gene duplication in strain EAN. Use of these techniques on the Thermotogales genome sequences revealed IS element bias and a clear instance of genome rearrangement mediated by IS elements. ^ While IS element activity is often determined by transposition into specialized plasmids, we provide evidence that suggests transcriptome sequencing may eventually be useful in quantifying transposition frequency. Three different conditions of Frankia sp. CcI3 cells were harvested and prepared into dscDNA libraries using an Illumina™ mRNA −seq kit. Comparison of the resultant sequencing data from those libraries suggests that nutrient deprivation is likely to increase the transcription of transposases in culture. While transcription does not necessarily correlate with transposition, the increased activity of these transposase ORFs suggests a likelihood of activity that will be confirmed with future studies. ^