title: Multiplex pyrosequencing of two polymorphisms in DNA repair gene XRCC1.
abstract: DNA repair enzymes play a pivotal role in platinum-based chemotherapy. Within the gene encoding for the base excision repair enzyme XRCC1, several nonsynonymous polymorphisms have been identified. It has been shown that the Arg399Gln single-nucleotide polymorphism results in a polymorphic enzyme that is less capable of initiating DNA repair. We developed a multiplex pyrosequence assay to simultaneously detect two nonsynonymous polymorphisms within the XRCC1 gene. Both of these polymorphisms resulted in amino acid changes: G/A in codon 399 changes Arg into Gln, and deletion of A in the second position of codon 576 results in a stopcodon. We established the frequency of these mutations in 270 patients suffering from colorectal cancer. Allele frequencies of G in second position of codon 399 and A in the second position codon 576 are 61.1 and 99.6%, respectively, in these patients. This fast and reliable method allows for simultaneous detection of the infrequent mutant C or CT alleles instead of the A deletion at codon 576. The method may be used in pharmacogenetic studies of platinum-based chemotherapy.
authors.affiliation: Department of Clinical Pharmacy and Toxicology, Leiden University Medical Center, P.O. Box 9600, 2300 RA Leiden, The Netherlands. vanderstra@lumc.nl
authors.forename: Tahar
authors.initials: T
authors.lastname: van der Straaten
authors.forename: Dinemarie
authors.initials: D
authors.lastname: Kweekel
authors.forename: Marco
authors.initials: M
authors.lastname: Tiller
authors.forename: Judith
authors.initials: J
authors.lastname: Bogaartz
authors.forename: Henk-Jan
authors.initials: HJ
authors.lastname: Guchelaar
mesh_terms: D015179:Colorectal Neoplasms / Q000235:genetics*; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000235:genetics*; D005787:Gene Frequency; D020022:Genetic Predisposition to Disease; D005820:Genetic Testing; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide; D017422:Sequence Analysis, DNA / Q000379:methods*; D000076105:X-ray Repair Cross Complementing Protein 1
chemical_list: D004268:DNA-Binding Proteins; D000076105:X-ray Repair Cross Complementing Protein 1; C000619371:XRCC1 protein, human