Authors: Kakuturu V. N. Rao; Priscilla Wilkins Stevens; Jeff G. Hall; Victor Lyamichev; Bruce P. Neri; David M. Kelso

Source: Nucleic Acids Research, Volume 31, Number 11, 01 June 2003 , pp. e66-e66(1)

Publisher: Oxford University Press

Abstract:

Here we report proof-of-principle for a microsphere-based genotyping assay that detects single nucleotide polymorphisms (SNPs) directly from human genomic DNA samples. This assay is based on a structure-specific cleavage reaction that achieves single base discrimination with a 5-nuclease which recognizes a tripartite substrate formed upon hybridization of target DNA with probe and upstream oligonucleotides. The assay is simple with two easy steps: a cleavage reaction, which generates fluorescent signal on microsphere surfaces, followed by flow cytometry analysis of the microspheres. Genomic DNA samples were genotyped for the SNP in the Apolipoprotein E gene at amino acid position 158. The assay successfully scored wild type, heterozygous and homozygous mutants. To our knowledge, this is the first report of a solid-support assay for detection of SNPs directly from genomic DNA without PCR amplification of the target.

Document Type: Research article

DOI: http://dx.doi.org/10.1093/nar/gng066

Affiliations: 1: Third Wave Technologies, 502 South Rosa Road, Madison, WI 53719-1256, USA

Publication date: 2003-06-01

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• Nucleic Acids Research (NAR) is a fully Open Access journal, providing rapid publication of leading edge research into the nucleic acids under the following categories: chemistry, computational biology, genomics, molecular biology, nucleic acid enzymes, RNA and structural biology. There is a Survey and Summary section, and methods papers are published
  in NAR Methods Online. Each year the first issue is devoted to biological databases, and a later issue to relevant web-based software resources.

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