Breaks Labeling in situ and sequencing (BLISS)

Abstract Precisely measuring the location and frequency of DNA double-strand breaks \(DSBs) along the genome is instrumental to understanding genomic fragility, but current methods are limited in versatility, sensitivity, or practicality. Here, we present Breaks Labeling _In Situ_ and Sequencing \(BLISS), featuring: 1) direct labeling of DSBs in fixed cells or tissue sections on a solid surface; 2) low-input requirement by linear amplification of tagged DSBs by _in vitro_ transcription; 3) quantification of DSBs through unique molecular identifiers; and 4) easy scalability and multiplexing. We apply BLISS to profile endogenous and exogenous DSBs in low-input samples of cancer cells, embryonic stem cells, and liver tissue. We demonstrate the sensitivity of BLISS by assessing the genome-wide off-target activity of two CRISPR-associated RNA-guided endonucleases, Cas9 and Cpf1, observing that Cpf1 has higher specificity than Cas9. Our results establish BLISS as a versatile, sensitive, and efficient method for genome-wide DSB mapping in many applications. W.Yan and R.Mirzazadeh equally contributed to this work.