Title :

Optimization of next-generation sequencing technologies for von hippel lindau (vhl) mosaic mutation detection and development of confirmation methods

Author(s) :

Coppin, Lucie [Auteur]
Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer (JPArc) - U1172
Plouvier, Pascal [Auteur]
Crepin, Michel [Auteur]
Jourdain, Anne-Sophie [Auteur]
Ait-Yahya, Emilie [Auteur]
Richard, Stephane [Auteur]
Paillerets, Brigitte Bressac-De [Auteur]
Cardot-Bauters, Catherine [Auteur]
Lejeune, Sophie [Auteur]
Leclerc, Julie [Auteur]
Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer (JPArc) - U1172
Pigny, Pascal [Auteur]
Centre de Recherche Jean-Pierre AUBERT Neurosciences et Cancer (JPArc) - U1172

Journal title :

The Journal of molecular diagnostics . JMD

Abbreviated title :

J Mol Diagn

Publication date :

2019-02-04

ISSN :

1943-7811

HAL domain(s) :

Sciences du Vivant [q-bio]

English abstract : [en]

Von Hippel-Lindau disease (VHL) is a monogenic disorder characterized by the development of tumors affecting the central nervous system, kidney, pancreas, or adrenal glands, and due to germline mutations in the VHL tumor ...
Show more >Von Hippel-Lindau disease (VHL) is a monogenic disorder characterized by the development of tumors affecting the central nervous system, kidney, pancreas, or adrenal glands, and due to germline mutations in the VHL tumor suppressor gene. About 5% of patients with a typical VHL phenotype have no mutation detected by conventional techniques, so a postzygotic VHL mosaicism can be suspected. The aim of this study was therefore to implement a next-generation sequencing (NGS) strategy for VHL mosaic mutation detection, including an optimization of the original Personal Genome Machine design by enrichment with oligonucleotides corresponding to amplicons with insufficient depth of coverage. Two complementary strategies were developed for the confirmation of mosaic mutations identified by NGS, SNaPshot for variants present at an allelic ratio greater than 5%, and droplet digital PCR for allelic ratio above 1%. VHL mutant plasmids were generated to assess VHL mosaic mutation detection in different exons and to set up an internal quality control that could be included in each run or regularly to validate the assay. This strategy was applied to 47 patients with a suggestive or clinical VHL disease, and mosaic mutations were identified in 8.5% of patients. In conclusion, NGS technologies combined with SNaPshot or droplet digital PCR allow the detection and confirmation of mosaic mutations in a clinical laboratory setting.Show less >

Language :

Anglais

Audience :

Internationale

Popular science :

Non

Administrative institution(s) :

CNRS
Université de Lille

Collections :

• Maladies Rares du Développement : Génétique, Régulation et Protéomique (RADEME) - ULR 7364
• Miniaturisation pour la Synthèse, l'Analyse et la Protéomique (MSAP) - USR 3290

Submission date :

2021-09-02T07:01:23Z