A Reliable Targeted Next-Generation Sequencing Strategy for Diagnosis of Myopathies and Muscular Dystrophies, Especially for the Giant Titin and Nebulin Genes

Abstract : Myopathies and muscular dystrophies (M-MDs) are genetically heterogeneous diseases, with more than 100 identified genes, including the giant and complex titin (TTN) and nebulin (NEB) genes. Next-generation sequencing technology revolutionized M-MD diagnosis and revealed high frequency of TTN and NEB variants. We developed a next-generation sequencing diagnostic strategy targeted on the coding sequences of 135 M-MD genes. Comparison of two targeted capture technologies (SeqCap EZ Choice library capture kit (Roche-Nimblegen) and Nextera Rapid Capture Custom Enrichment kit (Illumina)) and of two whole exome sequencing kits (SureSelect V5 (Agilent) and TruSeq RapidExome capture (Illumina)) revealed best coverage with the SeqCap EZ Choice protocol. A marked decrease in coverage was observed with the other kits, affecting mostly the first exons of genes and the repeated regions of TTN and NEB. Bioinformatics analysis strategy was fine-tuned to achieve optimal detection of variants, including small insertions/deletions (INDELs) and Copy Number Variants (CNVs). Analysis of a cohort of 128 patients allowed the detection of 52 substitutions, 13 INDELs (including a trinucleotide repeat expansion) and three CNVs. Two INDELs were localized in the repeated regions of NEB, suggesting that these mutations may be frequent but underestimated. A large deletion was also identified in TTN that is, to our knowledge, the first published CNV in this gene.
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https://hal.umontpellier.fr/hal-01800470
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Soumis le : vendredi 24 janvier 2020 - 09:07:23
Dernière modification le : samedi 25 janvier 2020 - 01:01:27

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Reda Zenagui, Delphine Lacourt, Henri Pégeot, Kevin Yauy, Raul Juntas Morales, et al.. A Reliable Targeted Next-Generation Sequencing Strategy for Diagnosis of Myopathies and Muscular Dystrophies, Especially for the Giant Titin and Nebulin Genes. The Journal of Molecular Diagnostics, Elsever, 2018, 20 (4), pp.533-549. ⟨10.1016/j.jmoldx.2018.04.001⟩. ⟨hal-01800470⟩

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