Genomewide nucleotide-resolution mapping of single-strand breaks and lesions by GLOE-Seq [Human_Ligase_inactivation]

Numerous methods are available for the mapping of DNA lesions ranging from double-strand breaks (DSBs) to incorporated ribonucleotides. We now present a technology based on the Genomewide Ligation of 3'-OH Ends (GLOE-Seq) and an associated computational pipeline designed for single-stranded breaks (SSBs), but versatile enough to be applied to any lesion that is convertible into a free 3'-OH terminus. We demonstrate its applicability to the mapping of Okazaki fragments without prior size selection and detect biases and asymmetries in the distribution of spontaneous SSBs in budding yeast and human DNA. Overall design: V. Analysis of Okazaki fragments and global DNA single-strand breaks in human cells DNA ligase-competent and -compromised HCT116 cells were subjected to GLOE-Seq analysis in two replicates each. Ligase-competent cells were used to map the genomewide distribution of DNA single-strand breaks. Cells with inactivated DNA Ligase 1 and 3 were used to map the distribution of Okazaki fragments.