Genome-wide strand-specific mapping of cleavage sites induced by topoisomerase I in E. coli

Cleavage sites induced by the EcTopoI G116S M320V topoisomerase mutant were identified in E. coli genome with a single-nucleotide resolution using Topo-Seq. Expression of EcTopoI G116S M320V strepII-tagged mutant (“intrinsically-poisoned” mutant known to form stable covalent complexes with DNA) was transiently (30 min) induced from a plasmid. Covalent complexes with DNA were enriched by immunoprecipitation and obtained DNA fragments were sequenced. Non-induced samples and samples without immunoprecipitation step served as controls. 3 biological replicates were processed and sequenced.