Hybridization Capture-Based Next-Generation Sequencing to Evaluate Coding Sequence and Deep Intronic Mutations in the NF1 Gene
Neurofibromatosis 1 (NF1) is one of the most common genetic disorders and is caused by mutations in the NF1 gene. NF1 gene mutational analysis presents a considerable challenge because of its large size, existence of highly homologous pseudogenes located throughout the human genome, absence of mutat...
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doaj-68a107b5b5604b66adb99308deb042e12020-11-25T00:30:58ZengMDPI AGGenes2073-44252016-12-0171213310.3390/genes7120133genes7120133Hybridization Capture-Based Next-Generation Sequencing to Evaluate Coding Sequence and Deep Intronic Mutations in the NF1 GeneKarin Soares Cunha0Nathalia Silva Oliveira1Anna Karoline Fausto2Carolina Cruz de Souza3Audrey Gros4Thomas Bandres5Yamina Idrissi6Jean-Philippe Merlio7Rodrigo Soares de Moura Neto8Rosane Silva9Mauro Geller10David Cappellen11Graduate Program in Pathology, School of Medicine, Universidade Federal Fluminense, Niterói 24033-900, BrazilAnatomy Pathology Service, Hospital Universitário Antônio Pedro, Universidade Federal Fluminense, Niterói 24033-900, BrazilAnatomy Pathology Service, Hospital Universitário Antônio Pedro, Universidade Federal Fluminense, Niterói 24033-900, BrazilSchool of Biomedicine, Universidade Federal Fluminense, Niterói 24210-130, BrazilService de Biologie des Tumeurs, Centre Hospitalier Universitaire de Bordeaux, Hôpital du Haut Lévêque, Pessac F-33604, FranceService de Biologie des Tumeurs, Centre Hospitalier Universitaire de Bordeaux, Hôpital du Haut Lévêque, Pessac F-33604, FranceInserm (Institut National de la Santé et de la Recherche Médicale) U1053, Bordeaux Research in Translational Oncology (BaRITON) and University of Bordeaux, Bordeaux F-33076, FranceService de Biologie des Tumeurs, Centre Hospitalier Universitaire de Bordeaux, Hôpital du Haut Lévêque, Pessac F-33604, FranceBiology Institute, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-599, BrazilCarlos Chagas Filho Biophysics Institute, Universidade Federal do Rio de Janeiro, Rio de Janeiro 21941-599, BrazilDepartment of Immunology and Microbiology, School of Medicine, Centro Universitário Serra dos Órgãos, Teresópolis 25964-004, BrazilService de Biologie des Tumeurs, Centre Hospitalier Universitaire de Bordeaux, Hôpital du Haut Lévêque, Pessac F-33604, FranceNeurofibromatosis 1 (NF1) is one of the most common genetic disorders and is caused by mutations in the NF1 gene. NF1 gene mutational analysis presents a considerable challenge because of its large size, existence of highly homologous pseudogenes located throughout the human genome, absence of mutational hotspots, and diversity of mutations types, including deep intronic splicing mutations. We aimed to evaluate the use of hybridization capture-based next-generation sequencing to screen coding and noncoding NF1 regions. Hybridization capture-based next-generation sequencing, with genomic DNA as starting material, was used to sequence the whole NF1 gene (exons and introns) from 11 unrelated individuals and 1 relative, who all had NF1. All of them met the NF1 clinical diagnostic criteria. We showed a mutation detection rate of 91% (10 out of 11). We identified eight recurrent and two novel mutations, which were all confirmed by Sanger methodology. In the Sanger sequencing confirmation, we also included another three relatives with NF1. Splicing alterations accounted for 50% of the mutations. One of them was caused by a deep intronic mutation (c.1260 + 1604A > G). Frameshift truncation and missense mutations corresponded to 30% and 20% of the pathogenic variants, respectively. In conclusion, we show the use of a simple and fast approach to screen, at once, the entire NF1 gene (exons and introns) for different types of pathogenic variations, including the deep intronic splicing mutations.http://www.mdpi.com/2073-4425/7/12/133Neurofibromatosis 1NF1 genenext generation sequencing |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Karin Soares Cunha Nathalia Silva Oliveira Anna Karoline Fausto Carolina Cruz de Souza Audrey Gros Thomas Bandres Yamina Idrissi Jean-Philippe Merlio Rodrigo Soares de Moura Neto Rosane Silva Mauro Geller David Cappellen |
spellingShingle |
Karin Soares Cunha Nathalia Silva Oliveira Anna Karoline Fausto Carolina Cruz de Souza Audrey Gros Thomas Bandres Yamina Idrissi Jean-Philippe Merlio Rodrigo Soares de Moura Neto Rosane Silva Mauro Geller David Cappellen Hybridization Capture-Based Next-Generation Sequencing to Evaluate Coding Sequence and Deep Intronic Mutations in the NF1 Gene Genes Neurofibromatosis 1 NF1 gene next generation sequencing |
author_facet |
Karin Soares Cunha Nathalia Silva Oliveira Anna Karoline Fausto Carolina Cruz de Souza Audrey Gros Thomas Bandres Yamina Idrissi Jean-Philippe Merlio Rodrigo Soares de Moura Neto Rosane Silva Mauro Geller David Cappellen |
author_sort |
Karin Soares Cunha |
title |
Hybridization Capture-Based Next-Generation Sequencing to Evaluate Coding Sequence and Deep Intronic Mutations in the NF1 Gene |
title_short |
Hybridization Capture-Based Next-Generation Sequencing to Evaluate Coding Sequence and Deep Intronic Mutations in the NF1 Gene |
title_full |
Hybridization Capture-Based Next-Generation Sequencing to Evaluate Coding Sequence and Deep Intronic Mutations in the NF1 Gene |
title_fullStr |
Hybridization Capture-Based Next-Generation Sequencing to Evaluate Coding Sequence and Deep Intronic Mutations in the NF1 Gene |
title_full_unstemmed |
Hybridization Capture-Based Next-Generation Sequencing to Evaluate Coding Sequence and Deep Intronic Mutations in the NF1 Gene |
title_sort |
hybridization capture-based next-generation sequencing to evaluate coding sequence and deep intronic mutations in the nf1 gene |
publisher |
MDPI AG |
series |
Genes |
issn |
2073-4425 |
publishDate |
2016-12-01 |
description |
Neurofibromatosis 1 (NF1) is one of the most common genetic disorders and is caused by mutations in the NF1 gene. NF1 gene mutational analysis presents a considerable challenge because of its large size, existence of highly homologous pseudogenes located throughout the human genome, absence of mutational hotspots, and diversity of mutations types, including deep intronic splicing mutations. We aimed to evaluate the use of hybridization capture-based next-generation sequencing to screen coding and noncoding NF1 regions. Hybridization capture-based next-generation sequencing, with genomic DNA as starting material, was used to sequence the whole NF1 gene (exons and introns) from 11 unrelated individuals and 1 relative, who all had NF1. All of them met the NF1 clinical diagnostic criteria. We showed a mutation detection rate of 91% (10 out of 11). We identified eight recurrent and two novel mutations, which were all confirmed by Sanger methodology. In the Sanger sequencing confirmation, we also included another three relatives with NF1. Splicing alterations accounted for 50% of the mutations. One of them was caused by a deep intronic mutation (c.1260 + 1604A > G). Frameshift truncation and missense mutations corresponded to 30% and 20% of the pathogenic variants, respectively. In conclusion, we show the use of a simple and fast approach to screen, at once, the entire NF1 gene (exons and introns) for different types of pathogenic variations, including the deep intronic splicing mutations. |
topic |
Neurofibromatosis 1 NF1 gene next generation sequencing |
url |
http://www.mdpi.com/2073-4425/7/12/133 |
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