Urological Research

, Volume 36, Issue 6, pp 297–302

Modification of primers for GRHPR genotyping: avoiding allele dropout by single nucleotide polymorphisms and homology sequence

  • Naohisa Takaoka
  • Tatsuya Takayama
  • Miki Miyazaki
  • Masao Nagata
  • Seiichiro Ozono
Symposium Paper

DOI: 10.1007/s00240-008-0159-z

Cite this article as:
Takaoka, N., Takayama, T., Miyazaki, M. et al. Urol Res (2008) 36: 297. doi:10.1007/s00240-008-0159-z

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.

Keywords

Single nucleotide polymorphism Allele dropout Glyoxylate/hydroxypyruvate reductase Genotyping Haplotype Linkage disequilibrium 

Copyright information

© Springer-Verlag 2008

Authors and Affiliations

  • Naohisa Takaoka
    • 1
  • Tatsuya Takayama
    • 1
  • Miki Miyazaki
    • 1
  • Masao Nagata
    • 1
  • Seiichiro Ozono
    • 1
  1. 1.Department of UrologyHamamatsu University School of MedicineHamamatsuJapan