Technical note: DNA typing for ovine MHC DRB1 using Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP)

Abstract

The ovine major histocompatibilty complex (Ovar) class II DRB1 second exon was amplified by polymerase chain reaction (PCR) from DNA samples of 52 Suffolk sheep. Polymerase chain reaction products were characterized by the restriction fragment length polymorphism (RFLP) technique using nine restriction enzymes, RsaI, HaeIII, SacI, SacII, DdeI, NciI, Hin1I, EcoRI, and BstNI, yielding 13 types. Sequencing of cloned PCR products identified 16 Ovar-DRB1 alleles. Collectively, all PCR-RFLP patterns exactly matched those predicted from DNA sequences. These findings strongly indicate that the PCR-RFLP method using a combination of nine restriction endonucleases is a very powerful tool in Ovar typing.