Implementation and Data Analysis of Tn-seq, Whole-Genome Resequencing, and Single-Molecule Real-Time Sequencing for Bacterial Genetics

Abstract

ABSTRACT Few discoveries have been more transformative to the biological sciences than the development of DNA sequencing technologies. The rapid advancement of sequencing and bioinformatics tools has revolutionized bacterial genetics, deepening our understanding of model and clinically relevant organisms. Although application of newer sequencing technologies to studies in bacterial genetics is increasing, the implementation of DNA sequencing technologies and development of the bioinformatics tools required for analyzing the large data sets generated remain a challenge for many. In this minireview, we have chosen to summarize three sequencing approaches that are particularly useful for bacterial genetics. We provide resources for scientists new to and interested in their application. Here, we discuss the analysis of data from transposon mutagenesis followed by deep sequencing (Tn-seq) to determine gene disruptions differentially represented in a mutant population and Illumina sequencing for identification of suppressor or other mutations, and we summarize single-molecule real-time (SMRT) sequencing for de novo genome assembly and the use of the output data for detection of DNA base modifications.