High speed single nucleotide polymorphism typing of a hereditary haemochromatosis mutation with capillary array electrophoresis microplates

Electrophoresis. 2000 Jul;21(12):2352-8. doi: 10.1002/1522-2683(20000701)21:12<2352::AID-ELPS2352>3.0.CO;2-G.

Abstract

A single nucleotide polymorphism (SNP) typing assay is developed and evaluated on a microfabricated capillary array electrophoresis system. Using fluorescently labeled allele-specific primers, the S65C (193A-->T) substitution associated with hereditary haemochromatosis in the HFE gene is genotyped. The covalently labeled polymerase chain reaction (PCR) products are separated on a microfabricated radial capillary array electrophoresis microplate using nondenaturing gel media in under two minutes. Detection is accomplished with a laser-excited rotary confocal scanner. The Rox-labeled A-allele specific amplicon (211 bp) is differentiated from the R110-labeled T-allele specific amplicon (201 bp) by both size and color. This study demonstrates the feasibility of using allele-specific PCR with covalently labeled primers for high speed fluorescent SNP typing on microfabricated radial capillary array electrophoresis microplates.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Alleles
  • Electrophoresis, Capillary / methods*
  • Genes, MHC Class I
  • Genetic Diseases, Inborn / genetics
  • Genotype
  • HLA Antigens / genetics*
  • Hemochromatosis / genetics*
  • Hemochromatosis Protein
  • Histocompatibility Antigens Class I / genetics*
  • Humans
  • Membrane Proteins*
  • Point Mutation
  • Polymerase Chain Reaction / methods
  • Polymorphism, Single Nucleotide*
  • Time Factors

Substances

  • HFE protein, human
  • HLA Antigens
  • Hemochromatosis Protein
  • Histocompatibility Antigens Class I
  • Membrane Proteins