Identification of RNA-DNA Hybrids Associated with R-Loops at the IgH Switch Sequence in Activated B Cells

During transcription and replication, R-loops that contain RNA-DNA hybrids are generated across numerous genomic loci and contribute to many biological events. Using S9.6, a monoclonal antibody against RNA-DNA hybrids, accelerated the study of R-loop biology. An outpouring of recent studies has implicated various contributions of R-loop in physiological cellular functions. Earlier studies using nondenaturing sodium bisulfite probing also supported existence of DNA-RNA hybrids formation in mammalian cells. In activated B cells, RNA-DNA hybrids formation at IgH gene locus of B cells is crucial for class switch recombination that ensure the proper effector function of the antibody. Here, we describe the identification of R-loops associated with the IgH locus using RNA-DNA hybrid immunoprecipitation sequencing and nondenaturing sodium bisulfite probing. This will be helpful for future studies of R-loops status on whole genome as well as on IgH locus in B cells.