Project description:The goal of this study was to compare the qualitative and quantitative differences in the presence of DNA:RNA hybrids (R-loops) between human wild type and PrimPol knockout human iPS cell lines. Our initial genetic experiments on a reporter locus have showed a link between the formation of R-loop and non-B DNA motifs; we have therefore analysed the correlation between R-loop abundance and of sequences computationaly predicted to form triplex-DNA or G-quadruplex forming sequences, and whether this changes between the two cell lines.
Project description:G-quadruplex (G4) are four‑stranded DNA secondary structures that form in guanine‑rich regions of the genome, which can enhance or repress gene expression. An R-loop is a special triple-stranded nucleic acid structure formed when nascent RNA invades double-stranded DNA (dsDNA) during transcription. G-loops are constituted by one or more DNA G4 on one strand and a stable RNA/DNA hybrid on the other. We developed the HepG4-seq for mapping the native G4s and the HBD-seq for mapping native R-loops. We combined the HepG4-seq and HBD-seq to profile the genomic native G-loops, which are regions co-occupied by both native G4s and R-loops, in both HEK293 cells and mouse embryonic stem cells (mESCs).
Project description:The goal of this study was to compare the qualitative and quantitative differences in the presence of DNA:RNA hybrids (R-loops) between chicken DT40 wild type and PrimPol knockout cells. Our initial genetic experiments on a reporter locus have showed a link between the formation of R-loop and non-B DNA motifs; we have therefore analysed the correlation between R-loop abundance and of sequences computationaly predicted to form triplex-DNA or G-quadruplex forming sequences, and whether this changes between the two cell lines.
Project description:Distribution of R-loops on genomic sites was studied for exponentially growing Escherichia coli in different conditions using strand-specific DRIP-Seq with S9.6 antibodies.
Project description:R-loops represent an abundant class of large non-B DNA structures in genomes. Even though they form transiently and at modest frequencies, interfering with R-loop formation or dissolution significantly impacts genome stability. Addressing the mechanism(s) of R-loop-mediated genome destabilization requires a precise characterization of their distribution in genomes. A number of independent methods have been developed to visualize and map R-loops, but their results are at times discordant, leading to confusion. Here we review the main existing methodologies underlying R-loop mapping and assess their limitations and the robustness of existing datasets. We offer a set of best practices to improve the reproducibility of maps, hoping that such guidelines could be useful for authors and referees alike. Finally, we offer a possible resolution to the apparent contradictions in R-loop mapping outcomes between antibody-based and RNase H1-based mapping approaches.