title: Identification of mutations in the repeated part of the autosomal dominant polycystic kidney disease type 1 gene, PKD1, by long-range PCR.
abstract: We have used long-range PCR to identify mutations in the duplicated part of the PKD1 gene. By means of a PKD1-specific primer in intron 1, an approximately 13.6-kb PCR product that includes exons 2-15 of the PKD1 gene has been used to search for mutations, by direct sequence analysis. This region contains the majority of the predicted extracellular domains of the PKD1-gene product, polycystin, including the 16 novel PKD domains that have similarity to immunoglobulin-like domains found in many cell-adhesion molecules and cell-surface receptors. Direct sequence analysis of exons encoding all the 16 PKD domains was performed on PCR products from a group of 24 unrelated patients with autosomal dominant polycystic kidney disease (ADPKD [MIM 173900]). Seven novel mutations were found in a screening of 42% of the PKD1-coding region in each patient, representing a 29% detection rate; these mutations included two deletions (one of 3 kb and the other of 28 bp), one single-base insertion, and four nucleotide substitutions (one splice site, one nonsense, and two missense). Five of these mutations would be predicted to cause a prematurely truncated protein. Two coding and 18 silent polymorphisms were also found. When, for the PKD1 gene, this method is coupled with existing mutation-detection methods, virtually the whole of this large, complex gene can now be screened for mutations.
authors.affiliation: Departments of Medical Genetics, Addenbrooke's Hospital, Cambridge, United Kingdom.
authors.forename: R
authors.initials: R
authors.lastname: Thomas
authors.forename: R
authors.initials: R
authors.lastname: McConnell
authors.forename: J
authors.initials: J
authors.lastname: Whittacker
authors.forename: P
authors.initials: P
authors.lastname: Kirkpatrick
authors.forename: J
authors.initials: J
authors.lastname: Bradley
authors.forename: R
authors.initials: R
authors.lastname: Sandford
mesh_terms: D001483:Base Sequence; D005091:Exons; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D016891:Polycystic Kidney, Autosomal Dominant / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D011110:Polymorphism, Genetic; D017433:Protein Structure, Secondary; D011506:Proteins / Q000737:chemistry / Q000235:genetics*; D017422:Sequence Analysis, DNA / Q000379:methods; D050396:TRPP Cation Channels
chemical_list: D011506:Proteins; D050396:TRPP Cation Channels