Multi-primer target PCR for rapid identification of bovine DRB3 alleles

Multi-primer target polymerase chain reaction (MPT-PCR) is a rapid method f or the identification of specific BoLA-DRB3 alleles. In a single PCR reacti on, the presence of two alleles associated with increased risk, DRB3.2*23 ( DRB3*2701-2703, 2705-2707) and decreased risk, DRB3.2*16 (DRB3*1501, 1502), of mastitis in Canadian Holstein can be detected. Two outer primers amplif y exon 2 of DRB3. Simultaneously, two inner, allele-specific primers amplif y individual alleles. Initially, 40 cows previously typed by PCR-restrictio n fragment length polymorphism (PCR-RFLP) were genotyped using the multi-pr imer approach. An additional 30 cows were first genotyped by multi-primer t arget PCR, then by PCR-RFLP. All animals were correctly identified and ther e were no false positives. This technique can readily be modified to identi fy other BoLA alleles of interest.