Molecular Genetic Analysis of Patients with Duchenne/Becker Muscular Dystrophy by Multiplex Ligation-Dependent Probe Amplification and Next-Generation Sequencing Techniques

Abstract

Background: Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) are neuromuscular genetic disorders, which are characterized by mutations in the dystrophin gene. Large deletions or duplications in the dystrophin gene are found in approximately 60 to 70% of cases, and the remaining have point mutations, small deletions, or insertions. Objectives: The aim of this study was to assess mutations in the dystrophin gene in clinically suspicious subjects of DMD/ BMD by multiplex ligation-dependent probe amplification (MLPA) and next-generation sequencing (NGS) techniques. Methods: This study consisted of 13 clinically suspected DMD/BMD patients. MLPA, NGS, and Sanger sequencing were used to determine mutations in the dystrophin gene. In order to minimize the time and cost, MLPA (SALSA P034/P035 DMD test kit) was performed as the first step to detect large deletions or duplications, followed by NGS and Sanger sequencing for MLPA-negative cases. Results: This study included nine males and four females with clinical suspicion of DMD or BMD. The MLPA was performed for all cases, among which ten cases were definitely diagnosed with DMD/BMD. NGS and Sanger sequencing were performed for the three remaining subjects to find out possible point mutations and small deletions, which could not be detected by MLPA. Nine intragenic deletions were identified, mostly in exons 46 to 47 and 4 to 7. Furthermore, an intragenic triplication was detected in exon 71 by MLPA. Conclusions: In agreement with other reports, our findings indicated that exonic deletions of the dystrophin gene are more frequent than duplications and point mutations in Duchenne/ Becker muscular dystrophy. Most deletions were demonstrated to occur in exons 46 to 47 and 4 to 7.