Approaches for Mapping and Analysis of R‐loops

Abstract R‐loops are nucleic acid structures composed of a DNA:RNA hybrid with a displaced non‐template single‐stranded DNA. Current approaches to identify and map R‐loop formation across the genome employ either an antibody targeted against R‐loops (S9.6) or a catalytically inactivated form of RNase H1 (dRNH1), a nuclease that can bind and resolve DNA:RNA hybrids via RNA exonuclease activity. This overview article outlines several ways to map R‐loops using either methodology, explaining the differences and similarities among the approaches. Bioinformatic analysis of R‐loops involves several layers of quality control and processing before visualizing the data. This article provides resources and tools that can be used to accurately process R‐loop mapping data and explains the advantages and disadvantages of the resources as compared to one another. © 2024 Wiley Periodicals LLC.