title: Targeted sequencing of a pediatric metabolic bone gene panel using a desktop semiconductor next-generation sequencer.
abstract: Metabolic bone disorders in children frequently are heritable, but the expanding number of genes associated with these conditions makes it difficult to perform molecular diagnosis. In the present study, we therefore evaluated a semiconductor (SC)-based sequencing system for this purpose. A total of 65 DNA samples were analyzed comprising 24 samples from patients with 27 known pathogenic mutations, 6 samples from patients with prior negative Sanger sequencing, and 35 consecutive samples from patients with suspected heritable metabolic bone disorders who had not had prior molecular diagnosis. In the samples with known pathogenic mutations, 26 of 27 mutations were identified by SC sequencing. All single nucleotide variants were correctly identified, but a 7-nucleotide duplication in CYP27B1 was not detected. SC sequencing revealed two pathogenic mutations in the six samples where prior Sanger sequencing had failed to identify a mutation. Finally, pathogenic mutations were found in 27 samples of patients with unknown mutation status (15 in COL1A1, 9 in COL1A2, 1 in LEPRE1, 1 in LRP5, 1 in PHEX). Subsequent Sanger sequencing confirmed the mutations in all 27 samples. In conclusion, we found that SC sequencing is suitable for the diagnosis of heritable metabolic bone disorders in children.
authors.affiliation: Shriners Hospital for Children and McGill University, 1529 Cedar Avenue, Montreal, QC, H3G 1A6, Canada, frauch@shriners.mcgill.ca.
authors.forename: Frank
authors.initials: F
authors.lastname: Rauch
authors.forename: Liljana
authors.initials: L
authors.lastname: Lalic
authors.forename: Francis H
authors.initials: FH
authors.lastname: Glorieux
authors.forename: Pierre
authors.initials: P
authors.lastname: Moffatt
authors.forename: Peter
authors.initials: P
authors.lastname: Roughley
mesh_terms: D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase / Q000235:genetics; D001842:Bone and Bones / Q000378:metabolism*; D002648:Child; D024042:Collagen Type I / Q000235:genetics; D000090602:Collagen Type I, alpha 1 Chain; D004247:DNA / Q000235:genetics; D020022:Genetic Predisposition to Disease; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D060492:Low Density Lipoprotein Receptor-Related Protein-5 / Q000235:genetics; D008562:Membrane Glycoproteins / Q000235:genetics; D009154:Mutation; D053402:PHEX Phosphate Regulating Neutral Endopeptidase / Q000235:genetics; D010372:Pediatrics; D064828:Prolyl Hydroxylases; D011509:Proteoglycans / Q000235:genetics; D012666:Semiconductors; D017422:Sequence Analysis, DNA / Q000379:methods
chemical_list: D024042:Collagen Type I; D000090602:Collagen Type I, alpha 1 Chain; C555482:LRP5 protein, human; D060492:Low Density Lipoprotein Receptor-Related Protein-5; D008562:Membrane Glycoproteins; D011509:Proteoglycans; D004247:DNA; D064828:Prolyl Hydroxylases; C485657:P3H1 protein, human; D015090:25-Hydroxyvitamin D3 1-alpha-Hydroxylase; D053402:PHEX Phosphate Regulating Neutral Endopeptidase; C506231:PHEX protein, human