Nanopore assay for fingerprinting DNA binding and quantifying real-time cleavage by catalytically active Cas9 enzyme.

ABSTRACT: Nanopore sensing, a high-resolution DNA sequencing technology, is rapidly expanding into novel and exciting directions of probing specific DNA-enzyme interactions. Although proven excellent for the detecting structural features of bare DNA, quantitative measurements on enzyme-DNA complexes and their real-time activity are lagging and only starting to emerge for long DNA templates. Signal-to-noise requirement and high translocation speeds make it difficult to detect protein bound on biologically relevant plasmid-length DNA. To this end we report accurate position detection of a catalytically active Cas9 bound to its single or multiple target sites on the DNA. Protein position is fingerprinted using event charge deficit (ECD) based analysis of the high signal-to-noise electrical signals as t

SUBMITTER: Nagpure P 

PROVIDER: S-EPMC12639772 | biostudies-literature | 2025 Nov

REPOSITORIES: biostudies-literature