Simple protocol for sequence-specific detection of mixed-base nucleic acids using a smart probe with NABs

Abstract

A fluorescent smart probe (SP) was used to detect a mixed-base ribonucleic acids sequence. While the SP presents excellent sensitivity for the target, it gives subtle discrimination between the perfect target sequence and several mismatch sequences. Its sequence-specificity for the target was greatly enhanced by using nucleic acid blockers (NABs), which are unlabeled, non-fluorescent hairpin oligonucleotides that are perfectly complementary to those mismatch sequences. This approach is simple, feasible at room temperature, requires no amplification enzymes, and it is suitable for applications requiring routine nucleic acids sequence detection and quantification methods such as genetic testing and biomedical diagnostics.