PCR-RFLP-based DQA1 typing of rhesus monkeys: sequence analysis of a new allele

In spite of the widespread use of rhesus monkeys (Macaca mulatta) in biomedical research, MHC typing of this species is not yet routine. Since suitable antibodies are lacking, serological typing of Mamu-DQA1 is not feasible. We developed a typing protocol for MhcMamu-DQA1 from published sequences of the second exon of Mamu-DQA1. This protocol is based on the amplification of the second exon of Mamu-DQA1 with one specific primer pair followed by a "diagnostic" digestion of the PCR products with, at most, 5 different restriction endonucleases. This modified PCR-RFLP permits the rapid identification of 11 out of 13 Mamu-DQA1 alleles in homozygous and heterozygous combinations. The protocol was validated by cloning and sequencing the PCR-products of several animals of different geographic origin. In addition, an as yet unknown allele was detected by PCR-RFLP and was subsequently cloned and its nucleotide sequence determined. All examined sequences except the new allele were identical to those previously published. Therefore, we assume that many of the Mamu-DQA1 alleles of rhesus monkeys have been identified molecularly and that the typing technique presented here can reliably identify Mamu-DQA1 alleles.