Major histocompatibility complex genotyping with massively parallel pyrosequencing

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Abstract
Nonhuman primates are key preclinical models for infectious disease, vaccine development and transplantation research, but their use has been hampered by the complexity and diversity of their major histocompatibility complex (MHC) class I genotypes. Wiseman and his colleagues provide a cost-effective solution to this problem using a next-generation pyrosequencing approach to high-resolution MHC genotyping in various nonhuman primates, identifying both known and new MHC class I alleles. Major histocompatibility complex (MHC) genetics dictate adaptive cellular immune responses, making robust MHC genotyping methods essential for studies of infectious disease, vaccine development and transplantation. Nonhuman primates provide essential preclinical models for these areas of biomedical research. Unfortunately, given the unparalleled complexity of macaque MHCs, existing methodologies are inadequate for MHC typing of these key model animals. Here we use pyrosequencing of complementary DNA–PCR amplicons as a general approach to determine comprehensive MHC class I genotypes in nonhuman primates. More than 500 unique MHC class I sequences were resolved by sequence-based typing of rhesus, cynomolgus and pig-tailed macaques, nearly half of which have not been reported previously. The remarkable sensitivity of this approach in macaques demonstrates that pyrosequencing is viable for ultra–high-throughput MHC genotyping of primates, including humans.