Research Letters in Biochemistry
Volume 2009 (2009), Article ID 960560, 5 pages
doi:10.1155/2009/960560
Research Letter

Multiplex Detection and Genotyping of Point Mutations Involved in Charcot-Marie-Tooth Disease Using a Hairpin Microarray-Based Assay

Yasser Baaj,1 Corinne Magdelaine,1 Virginie Ubertelli,2 Christophe Valat,2 Yoanne Mousseau,1 Hao Qiu,1 Benoît Funalot,3 Jean-Michel Vallat,3 and Franck G. Sturtz1

1Department of Biochemistry and Molecular Genetics, University of Limoges, 87025 Limoges, France
2Serial Genetics, 5 rue Henri Desbruères, 91000 Evry, France
3Department of Neurology, CHU Dupuytren, 87042 Limoges, France

Received 15 February 2009; Accepted 28 April 2009

Academic Editor: Vito Turk

Copyright © 2009 Yasser Baaj et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

We previously developed a highly specific method for detecting SNPs with a microarray-based system using stem-loop probes. In this paper we demonstrate that coupling a multiplexing procedure with our microarray method is possible for the simultaneous detection and genotyping of four point mutations, in three different genes, involved in Charcot-Marie-Tooth disease. DNA from healthy individuals and patients was amplified, labeled with Cy3 by multiplex PCR; and hybridized to microarrays. Spot signal intensities were 18 to 74 times greater for perfect matches than for mismatched target sequences differing by a single nucleotide (discrimination ratio) for “homozygous” DNA from healthy individuals. “Heterozygous” mutant DNA samples gave signal intensity ratios close to 1 at the positions of the mutations as expected. Genotyping by this method was therefore reliable. This system now combines the principle of highly specific genotyping based on stem-loop structure probes with the advantages of multiplex analysis.