DNA Probes for Cas12a-Based Assay with Fluorescence Anisotropy Enhanced Due to Anchors and Salts.

ABSTRACT: CRISPR/Cas12a is a potent biosensing tool known for its high specificity in DNA analysis. Cas12a recognizes the target DNA and acquires nuclease activity toward single-stranded DNA (ssDNA) probes. We present a straightforward and versatile approach to transforming common Cas12a-cleavable DNA probes into enhancing tools for fluorescence anisotropy (FA) measurements. Our study involved investigating 13 ssDNA probes with linear and hairpin structures, each featuring fluorescein at one end and a rotation-slowing tool (anchor) at the other. All anchors induced FA changes compared to fluorescein, ranging from 24 to 110 mr. Significant FA increases (up to 180 mr) were obtained by adding divalent metal salts (Mg²⁺, Ca²⁺, Ba²⁺), which influenced the rigidity and com

SUBMITTER: Safenkova IV 

PROVIDER: S-EPMC10741848 | biostudies-literature | 2023 Dec

REPOSITORIES: biostudies-literature