Genome-wide detection of DNA double-strand breaks by in-suspension BLISS

Nat Protoc. 2020 Dec;15(12):3894-3941. doi: 10.1038/s41596-020-0397-2. Epub 2020 Nov 2.

Abstract

sBLISS (in-suspension breaks labeling in situ and sequencing) is a versatile and widely applicable method for identification of endogenous and induced DNA double-strand breaks (DSBs) in any cell type that can be brought into suspension. sBLISS provides genome-wide profiles of the most consequential DNA lesion implicated in a variety of pathological, but also physiological, processes. In sBLISS, after in situ labeling, DSB ends are linearly amplified, followed by next-generation sequencing and DSB landscape analysis. Here, we present a step-by-step experimental protocol for sBLISS, as well as a basic computational analysis. The main advantages of sBLISS are (i) the suspension setup, which renders the protocol user-friendly and easily scalable; (ii) the possibility of adapting it to a high-throughput or single-cell workflow; and (iii) its flexibility and its applicability to virtually every cell type, including patient-derived cells, organoids, and isolated nuclei. The wet-lab protocol can be completed in 1.5 weeks and is suitable for researchers with intermediate expertise in molecular biology and genomics. For the computational analyses, basic-to-intermediate bioinformatics expertise is required.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Cell Line
  • DNA Breaks, Double-Stranded*
  • Genomics / methods*
  • Suspensions

Substances

  • Suspensions