Comprehensive Mutation Analysis for Congenital Muscular Dystrophy: A Clinical PCR-Based Enrichment and Next-Generation Sequencing Panel
Open Access
- 11 January 2013
- journal article
- research article
- Published by Public Library of Science (PLoS) in PLOS ONE
- Vol. 8 (1), e53083
- https://doi.org/10.1371/journal.pone.0053083
Abstract
The congenital muscular dystrophies (CMDs) comprise a heterogeneous group of heritable muscle disorders with often difficult to interpret muscle pathology, making them challenging to diagnose. Serial Sanger sequencing of suspected CMD genes, while the current molecular diagnostic method of choice, can be slow and expensive. A comprehensive panel test for simultaneous screening of mutations in all known CMD-associated genes would be a more effective diagnostic strategy. Thus, the CMDs are a model disorder group for development and validation of next-generation sequencing (NGS) strategies for diagnostic and clinical care applications. Using a highly multiplexed PCR-based target enrichment method (RainDance) in conjunction with NGS, we performed mutation detection in all CMD genes of 26 samples and compared the results with Sanger sequencing. The RainDance NGS panel showed great consistency in coverage depth, on-target efficiency, versatility of mutation detection, and genotype concordance with Sanger sequencing, demonstrating the test's appropriateness for clinical use. Compared to single tests, a higher diagnostic yield was observed by panel implementation. The panel's limitation is the amplification failure of select gene-specific exons which require Sanger sequencing for test completion. Successful validation and application of the CMD NGS panel to improve the diagnostic yield in a clinical laboratory was shown.This publication has 33 references indexed in Scilit:
- Relative frequency of congenital muscular dystrophy subtypes: Analysis of the UK diagnostic service 2001–2008Neuromuscular Disorders, 2012
- Exome Sequencing Identifies WDR35 Variants Involved in Sensenbrenner SyndromeAmerican Journal of Human Genetics, 2010
- Massively parallel sequencing of ataxia genes after array-based enrichmentHuman Mutation, 2010
- Mutation screening in 86 known X-linked mental retardation genes by droplet-based multiplex PCR and massive parallel sequencingThe HUGO Journal, 2009
- Congenital muscular dystrophy. Part II: a review of pathogenesis and therapeutic perspectivesArquivos de Neuro-Psiquiatria, 2009
- Congenital muscular dystrophy. Part I: a review of phenotypical and diagnostic aspectsArquivos de Neuro-Psiquiatria, 2009
- Congenital muscular dystrophies: New aspects of an expanding group of disordersBiochimica et Biophysica Acta (BBA) - Molecular Basis of Disease, 2007
- The Congenital Muscular Dystrophies: Recent Advances and Molecular InsightsPediatric and Developmental Pathology, 2006
- Congenital Muscular Dystrophies and the Extracellular MatrixSeminars in Pediatric Neurology, 2006
- The congenital muscular dystrophies in 2004: a century of exciting progressNeuromuscular Disorders, 2004