title: Identification of Disease-Causing Mutations by Functional Complementation of Patient-Derived Fibroblast Cell Lines.
abstract: Diagnosis of mitochondrial disorders is still hampered by their phenotypic and genotypic heterogeneity. In many cases, exome sequencing, the state-of-the-art method for genetically diagnosing mitochondrial disease patients, does not allow direct identification of the disease-associated gene but rather results in a list of variants in candidate genes. Here, we present a method to validate the disease-causing variant based on functional complementation assays. First, cell lines expressing a wild-type cDNA of the candidate genes are generated by lentiviral infection of patient-derived fibroblasts. Next, oxidative phosphorylation is measured by the Seahorse XF analyzer to assess rescue efficiency.
authors.affiliation: Institute of Human Genetics, Helmholtz Zentrum München, Ingolstaedter Landstr. 1, 85764, Neuherberg, Germany.|Institute of Human Genetics, Technische Universität München, 81675, Munich, Germany.
authors.forename: Laura S
authors.initials: LS
authors.lastname: Kremer
authors.affiliation: Institute of Human Genetics, Helmholtz Zentrum München, Ingolstaedter Landstr. 1, 85764, Neuherberg, Germany. prokisch@helmholtz-muenchen.de.|Institute of Human Genetics, Technische Universität München, 81675, Munich, Germany. prokisch@helmholtz-muenchen.de.
authors.forename: Holger
authors.initials: H
authors.lastname: Prokisch
mesh_terms: D002460:Cell Line; D005347:Fibroblasts / Q000378:metabolism*; D005816:Genetic Complementation Test; D020022:Genetic Predisposition to Disease; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D028361:Mitochondrial Diseases / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D010101:Oxygen Consumption; D010957:Plasmids / Q000235:genetics; D013223:Statistics as Topic; D013312:Stress, Physiological; D014161:Transduction, Genetic
keywords: Disease-associated gene; Exome sequencing; Functional complementation; Mitochondrial disorders; Oxygen consumption rate