A polymerase chain reaction screening strategy for the promoter of the canine dystrophin gene
- PMID: 10490068
A polymerase chain reaction screening strategy for the promoter of the canine dystrophin gene
Abstract
Objective: To develop a polymerase chain reaction (PCR) strategy to screen the dystrophin promoter(s) in dogs with cardiac and skeletal myopathies.
Animals: 9 Doberman Pinschers, 1 Dalmation, and 1 Saint Bernard with dilated cardiomyopathy (DCM); 1 Irish Terrier with muscular dystrophy; and 2 dystrophin-deficient German Shorthaired Pointers (GSHP).
Procedure: For each of the 3 unique exons associated with the muscle (M), Purkinje (P), and cortical (C) promoters of the dystrophin gene, each first exon, and the M promoter plus its first exon, were amplified, cloned, and sequenced. The M dystrophin transcript was amplified by reverse transcriptase PCR from skeletal and cardiac muscle RNA of 1 Doberman Pinscher and from skeletal muscle RNA of 1 GSHP.
Results: The M, P, and C first exons were amplified from all dogs except the 2 GSHP, which had a deletion encompassing the entire M, P, and C dystrophin promoter region. The M transcript could not be amplified from muscles of the GSHP, but was amplified from skeletal and cardiac muscle of the Doberman Pinscher. Sequencing of the product representing the M promoter and its first exon revealed no differences between clinically normal dogs and the Doberman Pinscher with DCM.
Conclusions and clinical relevance: We have ruled out a major rearrangement of the dystrophin promoter region as the universal cause of DCM in Doberman Pinschers or of Irish Terrier myopathy. Use of the strategy identified a large deletion of this region in muscle from the GSHP.
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