Highly Sensitive Diagnosis of 43 Monogenic Forms of Diabetes or Obesity Through One-Step PCR-Based Enrichment in Combination With Next-Generation Sequencing

• DOI: 10.2337/dc13-0698
• Publication Date: 2013-09-17T04:35:56Z
• Authors: Amélie Bonnefond; Julien Philippe; Emmanuelle Durand; Jean Muller; Sadia Saeed; Muhammad Arslan; Rosa Martínez; Franck De Graeve; Véronique Dhennin; Iandry Rabearivelo; Michel Polak; Hélène Cavé; Luis Castaño; Martine Vaxillaire; Jean-Louis Mandel; Olivier Sand; Philippe Froguel
• Keywords: Sanger sequencing; Indel

OBJECTIVE Accurate etiological diagnosis of monogenic forms diabetes and obesity is useful as it can lead to marked improvements in patient care genetic counseling. Currently, molecular based on Sanger sequencing restricted only a few genes, this technology expensive, time-consuming, labor-intensive. High-throughput next-generation (NGS) provides an opportunity develop innovative cost-efficient methods for sensitive multigene screening. RESEARCH DESIGN AND METHODS We assessed new method PCR enrichment microdroplets (RainDance Technologies) NGS using the Illumina HiSeq2000 43 or obesity. Forty patients carrying known causal mutation those subtypes according diagnostic laboratories were blindly reanalyzed. RESULTS Except one variant, we reidentified all mutations each associated with almost-perfect targets (mean 98.6%). failed call highly complex indel, although identified dramatic drop coverage at locus. In three patients, detected other putatively deleterious effect addition reported by laboratories. CONCLUSIONS Our approach efficient means screening genes As cost time deliver results have been key barriers uncovering cause many undiagnosed cases likely exist, present methodology should be considered displaying features