Allele-Specific Holliday Junction Formation: A New Mechanism of Allelic Discrimination for SNP Scoring
Abstract
We report here a new mechanism for allelic discrimination—allele-specific Holliday Junction formation. The Holliday Junction (HJ) is a unique DNA structure that can be formed in a sequence-nonspecific manner by routine PCR. To cause the PCR-based HJ formation to occur in an allele-specific manner, the PCR primers are manipulated such that an extra mismatch next to a SNP of interest is introduced between a target and a reference amplicon and a GC-clamp is added. Based on this new mechanism, novel SNP genotyping methods were developed, including a homogeneous fluorescence polarization (FP) competition assay that requires neither labeled primers/probes nor expensive enzymes/substrates. Using this novel genotyping technology, we were able to convert >95% of SNP sequences into genotyping assays that work well under a universal set of assay conditions and achieved 100% accuracy in clinical samples.
Footnotes
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Article and publication are at http://www.genome.org/cgi/doi/10.1101/gr.997703.
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↵3 Corresponding author. E-MAIL qinghong{at}freshgene.com; FAX (925) 681-4012.
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- Accepted April 18, 2003.
- Received November 15, 2002.
- Cold Spring Harbor Laboratory Press
