Microarray-based mutation detection in the dystrophin gene.
Duchenne and Becker muscular dystrophies (DMD and BMD) are X-linked recessive neuromuscular disorders caused by mutations in the dystrophin gene affecting approximately 1 in 3,500 males. The human dystrophin gene spans>2,200 kb, or roughly 0.1% of the genome, and is composed of 79 exons. The mutational spectrum of disease-causing alleles, including exonic copy number variations (CNVs), is complex. Deletions account for approximately 65% of DMD mutations and 85% of BMD mutations. Duplications occur in approximately 6 to 10% of males with either DMD or BMD. The remaining 30 to 35% of mutations consist of small deletions, insertions, point mutations, or splicing mutations, most of which introduce a premature stop codon. Laboratory analysis of dystrophin can be used to confirm a clinical diagnosis of DMD, characterize the type of dystrophin mutation, and perform prenatal testing and carrier testing for females. Current dystrophin diagnostic assays involve a variety of methodologies, including multiplex PCR, Southern blot analysis, multiplex ligation-dependent probe amplification (MLPA), detection of virtually all mutations-SSCP (DOVAM-S), and single condition amplification/internal primer sequencing (SCAIP); however, these methods are time-consuming, laborious, and do not accurately detect duplication mutations in the dystrophin gene. Furthermore, carrier testing in females is often difficult when a related affected male is unavailable. Here we describe the development, design, validation, and implementation of a high-resolution comparative genomic hybridization (CGH) microarray-based approach capable of accurately detecting both deletions and duplications in the dystrophin gene. This assay can be readily adopted by clinical molecular testing laboratories and represents a rapid, cost-effective approach for screening a large gene, such as dystrophin.
Hegde
Madhuri R
MR
Department of Human Genetics, Emory University School of Medicine, Atlanta, Georgia 30322, USA. mhegde@genetics.emory.edu
Chin
Ephrem L H
EL
Mulle
Jennifer G
JG
Okou
David T
DT
Warren
Stephen T
ST
Zwick
Michael E
ME
D016189:Dystrophin / Q000235:genetics*; D005260:Female; D020440:Gene Duplication; D005838:Genotype; D006801:Humans; D008297:Male; D025202:Molecular Diagnostic Techniques / Q000379:methods; D020388:Muscular Dystrophy, Duchenne / Q000175:diagnosis*; D009154:Mutation; D009693:Nucleic Acid Hybridization / Q000379:methods*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D017384:Sequence Deletion
D016189:Dystrophin