Conformational Studies of Bacterial Chromosomes by High-Throughput Sequencing Methods

Abstract

The development of next-generation sequencing technologies has allowed the application of different methods dedicated to the study of DNA-protein interactions and chromosome conformation to entire bacterial genome. By combining these approaches, the role of various parameters and factors involved in gene expression and chromosome organization can be disclosed at the molecular level over the full genome. Here we describe two methods that profoundly revolutionized our vision of DNA-protein interactions and spatial organization of chromosomes. Chromosome conformation capture (3C) coupled to deep sequencing (3C-seq) enables studies of the genome-wide chromosome folding and its control by different parameters and structural factors. Chromatin immunoprecipitation (ChIP) followed by high-throughput DNA sequencing (ChIP-seq) revealed the extent and regulation of DNA-protein interactions in vivo and highlight the role of structural factors in the control of chromosome organization. In this chapter, we describe a detailed protocol of 3C-seq and ChIP-seq experiments that, when combined, allows the spatial study of the chromosome and the factors that promote specific folding. Data processing and analysis for both experiments are also discussed.

Keywords: 3C-seq; ChIP-seq; Chromosome conformation; Condensin; Nucleoid-associated proteins.