title: Modification of primers for GRHPR genotyping: avoiding allele dropout by single nucleotide polymorphisms and homology sequence.
abstract: Mutation of primer site for genotyping by polymerase chain reaction (PCR) may cause allele dropout and other genotyping failures. Primary hyperoxaluria type 2 (PH2) is a rare inherited disease caused by overproduction of endogenous oxalate due to mutations in the glyoxylate/hydroxypyruvate reductase (GRHPR) gene. Here, to avoid allele dropout and primer annealing to multiple sites, and given the discrepancy in intron length between GRHPR gene data, we updated the primers used in the sequence assay of the GRHPR gene. These redesigned primers show potential in reducing detection failure of GRHPR mutations. In addition, we performed a single nucleotide polymorphism (SNP) linkage analysis of the GRHPR gene using direct sequencing with PCR amplification of specific alleles (DS-PASA). Using this technique, we sequenced four common SNPs between intron E and exon 6, which show linkage disequilibrium (LD) consisting of three types of haplotypes, similar to data from the HapMap SNP database.
authors.affiliation: Department of Urology, Hamamatsu University School of Medicine, 1-20-1 Handayama, Higashi-ku, Hamamatsu, Shizuoka, 431-3192, Japan.
authors.forename: Naohisa
authors.initials: N
authors.lastname: Takaoka
authors.forename: Tatsuya
authors.initials: T
authors.lastname: Takayama
authors.forename: Miki
authors.initials: M
authors.lastname: Miyazaki
authors.forename: Masao
authors.initials: M
authors.lastname: Nagata
authors.forename: Seiichiro
authors.initials: S
authors.lastname: Ozono
mesh_terms: D000429:Alcohol Oxidoreductases / Q000235:genetics*; D000483:Alleles; D001483:Base Sequence / Q000235:genetics; D004247:DNA / Q000235:genetics; D017931:DNA Primers / Q000235:genetics*; D005091:Exons / Q000235:genetics; D005838:Genotype; D006801:Humans; D006960:Hyperoxaluria, Primary / Q000235:genetics; D007438:Introns / Q000235:genetics; D015810:Linkage Disequilibrium / Q000235:genetics; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid
chemical_list: D017931:DNA Primers; D004247:DNA; D000429:Alcohol Oxidoreductases; C020830:glyoxylate reductase