title: Diagnosis of autosomal dominant polycystic kidney disease using efficient PKD1 and PKD2 targeted next-generation sequencing.
abstract: Molecular diagnostics of autosomal dominant polycystic kidney disease (ADPKD) relies on mutation screening of PKD1 and PKD2, which is complicated by extensive allelic heterogeneity and the presence of six highly homologous sequences of PKD1. To date, specific sequencing of PKD1 requires laborious long-range amplifications. The high cost and long turnaround time of PKD1 and PKD2 mutation analysis using conventional techniques limits its widespread application in clinical settings. We performed targeted next-generation sequencing (NGS) of PKD1 and PKD2. Pooled barcoded DNA patient libraries were enriched by in-solution hybridization with PKD1 and PKD2 capture probes. Bioinformatics analysis was performed using an in-house developed pipeline. We validated the assay in a cohort of 36 patients with previously known PKD1 and PKD2 mutations and five control individuals. Then, we used the same assay and bioinformatics analysis in a discovery cohort of 12 uncharacterized patients. We detected 35 out of 36 known definitely, highly likely, and likely pathogenic mutations in the validation cohort, including two large deletions. In the discovery cohort, we detected 11 different pathogenic mutations in 10 out of 12 patients. This study demonstrates that laborious long-range PCRs of the repeated PKD1 region can be avoided by in-solution enrichment of PKD1 and PKD2 and NGS. This strategy significantly reduces the cost and time for simultaneous PKD1 and PKD2 sequence analysis, facilitating routine genetic diagnostics of ADPKD.
authors.affiliation: Genomics and Disease Group, Bioinformatics and Genomics Programme, Centre for Genomic Regulation (CRG) Dr. Aiguader 88, 08003 Barcelona, Catalonia, Spain ; Universitat Pompeu Fabra (UPF) Barcelona, Catalonia, Spain ; Institut Hospital del Mar d'Investigacions Mèdiques (IMIM) 08003 Barcelona, Catalonia, Spain ; CIBER in Epidemiology and Public Health (CIBERESP) Barcelona, Catalonia, Spain.
authors.forename: Daniel
authors.initials: D
authors.lastname: Trujillano
authors.affiliation: Molecular Biology Laboratory, Fundació Puigvert, Instituto de Investigaciones Biomédicas Sant Pau (IIB-Sant Pau), Universitat Autònoma de Barcelona REDinREN, Instituto de Investigación Carlos III, Barcelona, Catalonia, Spain ; Nephrology Department, Fundació Puigvert, Instituto de Investigaciones Biomédicas Sant Pau (IIB-Sant Pau), Universitat Autònoma de Barcelona REDinREN, Instituto de Investigación Carlos III, Barcelona, Catalonia, Spain.
authors.forename: Gemma
authors.initials: G
authors.lastname: Bullich
authors.affiliation: Genomic and Epigenomic Variation in Disease Group, Centre for Genomic Regulation (CRG) Dr. Aiguader 88, 08003 Barcelona, Catalonia, Spain ; Universitat Pompeu Fabra (UPF) Barcelona, Catalonia, Spain.
authors.forename: Stephan
authors.initials: S
authors.lastname: Ossowski
authors.affiliation: Nephrology Department, Fundació Puigvert, Instituto de Investigaciones Biomédicas Sant Pau (IIB-Sant Pau), Universitat Autònoma de Barcelona REDinREN, Instituto de Investigación Carlos III, Barcelona, Catalonia, Spain.
authors.forename: José
authors.initials: J
authors.lastname: Ballarín
authors.affiliation: Nephrology Department, Fundació Puigvert, Instituto de Investigaciones Biomédicas Sant Pau (IIB-Sant Pau), Universitat Autònoma de Barcelona REDinREN, Instituto de Investigación Carlos III, Barcelona, Catalonia, Spain.
authors.forename: Roser
authors.initials: R
authors.lastname: Torra
authors.affiliation: Genomics and Disease Group, Bioinformatics and Genomics Programme, Centre for Genomic Regulation (CRG) Dr. Aiguader 88, 08003 Barcelona, Catalonia, Spain ; Universitat Pompeu Fabra (UPF) Barcelona, Catalonia, Spain ; Institut Hospital del Mar d'Investigacions Mèdiques (IMIM) 08003 Barcelona, Catalonia, Spain ; CIBER in Epidemiology and Public Health (CIBERESP) Barcelona, Catalonia, Spain ; Dexeus Woman's Health, Hospital Universitary Quiron Dexeus Barcelona, Catalonia, Spain.
authors.forename: Xavier
authors.initials: X
authors.lastname: Estivill
authors.affiliation: Molecular Biology Laboratory, Fundació Puigvert, Instituto de Investigaciones Biomédicas Sant Pau (IIB-Sant Pau), Universitat Autònoma de Barcelona REDinREN, Instituto de Investigación Carlos III, Barcelona, Catalonia, Spain ; Nephrology Department, Fundació Puigvert, Instituto de Investigaciones Biomédicas Sant Pau (IIB-Sant Pau), Universitat Autònoma de Barcelona REDinREN, Instituto de Investigación Carlos III, Barcelona, Catalonia, Spain.
authors.forename: Elisabet
authors.initials: E
authors.lastname: Ars
keywords: Autosomal dominant polycystic kidney disease; genetic counseling; molecular diagnostics; targeted NGS