Simultaneous detection of multiple single nucleotide polymorphism by single-strand-specific nuclease and PNA probe

The combination of PNA (peptide nucleic acid) and single-strand-specific nuclease have been used for detection of single nucleotide polymorphisms (SNPs). When DNA is perfectly complementary to PNA, it is protected from digestion by the nuclease. If there exists a single-base mismatch between them, however, the DNA is completely digested. These differences are visualized by using 3,3'-diethylthiadicarbocyanine (DiSc2(5)), which changes its color from blue to purple upon binding to PNA/DNA hybrids. In terms of this methodology, homozygous and heterozygous SNPs in apoE gene have been successfully analyzed. Furthermore, the multiplex SNPs are simultaneously genotyped. This technique provides a simple, straightforward, facile, and visual genetic screening, with no need for expensive and complicated equipment.