Silver Nanoparticles with Dual-Recognition via CRISPR/dCas9 for SERS Identification of Two KRAS Mutations in Nucleic Acid Targets

Abstract

Specifically identifying one-base mismatched gene sequences is of significance for early diagnosis, prognosis, treatment, and evaluation. However, traditional polymerase chain reaction and next-generation sequencing methods require relatively long, carefully implemented analytical procedures. Here, we report a nanoconjugate that can synchronously discriminate two types of KRAS gene mutation due to the dual-recognition mechanism of the CRISPR/dCas system with the surface-enhanced Raman scattering (SERS) response output. This method applied two sets of CRISPR/dCas9 units to collect all KRAS genes via a magnetic force and the specific mutant sequence labeled through SERS nanotags, forming a dualclamping architecture. This sensing conjugate was sufficient to quantify the amount of the two different mutations simultaneously with a low crossreactivity. Consequently, we deem that this sensing assay satisfies the great characteristics of a genetic analytical tool with high specificity, accuracy, and simplicity, highlighting its potential as an alternative for screening multigene events and relevant genetic biomarkers.