One hundred twenty-one dystrophin point mutations detected from stored DNA samples by combinatorial denaturing high-performance liquid chromatography

Annalaura Torella, Amelia Trimarco, Francesca Del Vecchio Blanco, Anna Cuomo, Stefania Aurino, Giulio Piluso, Carlo Minetti, Luisa Politano, Vincenzo Nigro

Research output: Contribution to journalArticlepeer-review

Abstract

Duchenne and Becker muscular dystrophies are caused by a large number of different mutations in the dystrophin gene. Outside of the deletion/duplication "hot spots," small mutations occur at unpredictable positions. These account for about 15 to 20% of cases, with the major group being premature stop codons. When the affected male is deceased, carrier testing for family members and prenatal diagnosis become difficult and expensive. We tailored a cost-effective and reliable strategy to discover point mutations from stored DNA samples in the absence of a muscle biopsy. Samples were amplified in combinatorial pools and tested by denaturing high-performance liquid chromatography analysis. An anomalous elution profile belonging to two different pools univocally addressed the allelic variation to an unambiguous sample. Mutations were then detected by sequencing. We identified 121 mutations of 99 different types. Fifty-six patients show stop codons that represent the 46.3% of all cases. Three nonobvious single amino acid mutations were considered as causative. Our data support combinatorial denaturing high-performance liquid chromatography analysis as a clear-cut strategy for time and cost-effective identification of small mutations when only DNA is available.

Original languageEnglish
Pages (from-to)65-73
Number of pages9
JournalJournal of Molecular Diagnostics
Volume12
Issue number1
DOIs
Publication statusPublished - Jan 2010

ASJC Scopus subject areas

  • Molecular Medicine
  • Pathology and Forensic Medicine

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