Neutralized chimeric DNA probe for the improvement of GC-rich RNA detection specificity on the nanowire field-effect transistor

Silicon nanowire (SiNW) field-effect transistors (FETs) is a powerful tool in genetic molecule analysis because of their high sensitivity, short detection time, and label-free detection. In nucleic acid detection, GC-rich nucleic acid sequences form self- and cross-dimers and stem-loop structures, which can easily obtain data containing signals from nonspecific DNA binding. The features of GC-rich nucleic acid sequences cause inaccuracies in nucleic acid detection and hinder the development of precision medicine. To improve the inaccurate detection results, we used phosphate-methylated (neutral) nucleotides to synthesize the neutralized chimeric DNA oligomer probe. The probe fragment originated from a primer for the detection of hepatitis C virus (HCV) genotype 3b, and single-mismatched and perfect-matched targets were designed for single nucleotide polymorphisms (SNP) detection on the SiNW FET device. Experimental results revealed that the HCV-3b chimeric neutralized DNA (nDNA) probe exhibited better performance for SNP discrimination in 10 mM bis-tris propane buffer at 25 °C than a regular DNA probe. The SNP discrimination of the nDNA probe could be further improved at 40 °C on the FET device. Consequently, the neutralized chimeric DNA probe could successfully distinguish SNP in the detection of GC-rich target sequences under optimal operating conditions on the SiNW FET device.