Open AccessSNP analysis using catacleave probes
Author(s) -
Harvey John J.,
Brant Steven R.,
Knutson Jay R.,
Han Myun K.
Publication year - 2008
Publication title -
journal of clinical laboratory analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 50
eISSN - 1098-2825
pISSN - 0887-8013
DOI - 10.1002/jcla.20240
Subject(s) - snp , computational biology , biology , genetics , computer science , genotype , single nucleotide polymorphism , gene
CataCleave” probes are catalytically cleavable fluorescence probes having a chimeric deoxyribonucleic acid (DNA)‐ribonucleic acid (RNA)‐DNA structure that can be used for real‐time detection of single nucleotide polymorphisms (SNPs), insertions, and deletions. Fluorescent donor emission is normally quenched by Förster resonance energy transfer (FRET). Upon binding to the target, if the RNA/DNA hybrid is correctly base‐paired, ribonuclease (RNase) H will cleave the RNA moiety and the probe fragments will dissociate. FRET is lost and the donor fluorescence signal is recovered. A single‐base mismatch within the hybrid region causes probe cleavage to be significantly reduced. We designed CataCleave probes to detect SNPs located in the insulin‐like growth factor 2 (IGF‐2) gene and at position 702 within the NOD2/CARD15 gene. Probes were also designed to detect a six‐basepair deletion in the amelogenin gene and a partially methylated target DNA. Discrimination between wild‐type and SNP is demonstrated for both genes in homogeneous reactions under isothermal and temperature cycling conditions. These probes were also able to identify a multibase deletion and methylated DNA. Cleavage rates were proportional to target concentration. Probe length and position of fluorescent labels may also be modified for use in multiplexing high‐throughput SNP assays. This represents a novel method for the detection of SNPs. J. Clin. Lab. Anal. 22:192–203, 2008. © 2008 Wiley‐Liss, Inc.