Transposon mutagenesis libraries reveal novel molecular requirements during CRISPR RNA-guided DNA integration.

ABSTRACT: CRISPR-associated transposons (CASTs) direct DNA integration downstream of target sites using the RNA-guided DNA binding activity of nuclease-deficient CRISPR-Cas systems. Transposition relies on several key protein-protein and protein-DNA interactions, but little is known about the explicit sequence requirements governing efficient transposon DNA integration activity. Here, we exploit pooled library screening and high-throughput sequencing to reveal novel sequence determinants during transposition by the Type I-F Vibrio cholerae CAST system. On the donor DNA, large mutagenic libraries identified core binding sites recognized by the TnsB transposase, as well as an additional conserved region that encoded a consensus binding site for integration host factor (IHF). Remarkably, we foun

SUBMITTER: Walker MWG 

PROVIDER: S-EPMC9882353 | biostudies-literature | 2023 Jan

REPOSITORIES: biostudies-literature