High-fidelity amplified FISH for the detection and allelic discrimination of single mRNA molecules.

ABSTRACT: Amplification of signals by the hybridization chain reaction (HCR) is a powerful approach for increasing signal strength in single-molecule fluorescence in situ hybridization, but probes tagged with an HCR initiator sequence are prone to producing false signals. Here we describe a system of interacting hairpin binary probes in which the HCR initiator sequence is conditionally sequestered. The binding of these probes to a perfectly complementary target unmasks the initiator, enabling the generation of an amplified signal. This probe system can distinguish single-nucleotide variations within single mRNA molecules and produces amplified signals in situ for both mutant and wild-type variants, each in a distinguishable color. This technology will augment studies of imbalanced allelic expression

SUBMITTER: Marras SAE 

PROVIDER: S-EPMC6628817 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature