DNA conformation polymorphism analysis of DR52 associated HLA-DR antigens by polymerase chain reaction: a simple, economical and rapid examination for HLA matching in transplantation.

Abstract

HLA-DRB1 and -DRB3 alleles of DR52-associated (DR52ass) HLA-DR antigens were genotyped by a polymerase chain reaction (PCR) - based simple and practical method. Genomic DNAs from two hundred Japanese panels were subjected to PCR with two pairs of primers to separately amplify the DR52ass-DRB1 (DR3, 5, 6, and 8) alleles and DRB3 (DR52) alleles. The specific amplification revealed that 128 and 76 panels possessed DR52ass alleles and DRB3 alleles, respectively. PCR products from these panels were heat-denatured, electrophoresed in a non-denaturing polyacrylamide gel, and visualized by silver staining. Electrophoretic mobilities of the DNA samples were compared with those of the typing standards with known genotypes of DR52ass-DRB1 and DRB3 alleles. This method, designated PCR-DNA conformation polymorphism (DCP) analysis, allowed genotyping of the DR52ass-DRB1 and DRB3 alleles of panels without any sequence-specific oligonucleotide probe (SSOP) or restriction endonuclease, and the entire process after PCR could be completed within a few hours. Because the DR52ass-DRB1 and DRB3 alleles assigned by this method were shown to be identical to those determined by the PCR-SSOP method, PCR-DCP analysis was suggested to be a simple and practical HLA genotyping method.