SERS-melting: a new method for discriminating mutations in DNA sequences.
The reliable discrimination of mutations, single nucleotide polymorphisms (SNPs), and other differences in genomic sequence is an essential part of DNA diagnostics and forensics. It is commonly achieved using fluorescently labeled DNA probes and thermal gradients to distinguish between the matched and mismatched DNA. Here, we describe a novel method that uses surface enhanced (resonance) Raman spectroscopy (SER(R)S) to follow denaturation of dsDNA attached to a structured gold surface. This denaturation is driven either electrochemically or thermally on SERS active sphere segment void (SSV) gold substrates. Using this method, we can distinguish between wild type, a single point mutation (1653C/T), and a triple deletion (DeltaF 508) in the CFTR gene at the 0.02 attomole level, and the method can be used to differentiate the unpurified PCR products of the wild type and DeltaF 508 mutation. Our method has the potential to provide small, rapid, sensitive, reproducible platforms for detecting genetic variations and sequencing genes.
Mahajan
Sumeet
S
School of Chemistry, University of Southampton, Southampton SO17 1BJ, United Kingdom.
Richardson
James
J
Brown
Tom
T
Bartlett
Philip N
PN
D001483:Base Sequence; D004247:DNA / Q000737:chemistry* / Q000235:genetics*; D004220:Disulfides / Q000737:chemistry; D004563:Electrochemistry; D008855:Microscopy, Electron, Scanning; D009154:Mutation / Q000235:genetics; D015203:Reproducibility of Results; D013059:Spectrum Analysis, Raman / Q000379:methods*; D013499:Surface Properties; D044366:Transition Temperature
D004220:Disulfides; D004247:DNA