Polymorphisms of the equine major histocompatibility complex class II DRA locus J.J. Brown W. Thomson P. Clegg S. Eyre L.J. Kennedy J. Matthews S. Carter W.E.R. Ollier Authors’ affiliations: J.J. Brown 1,2 , W. Thomson 2 , P. Clegg 1 , S. Eyre 2 , L.J. Kennedy 2 , J. Matthews 1 , S. Carter 1 , W.E.R. Ollier 2 1 Department of Veterinary Clinical Science, University of Liverpool, South Wirral, UK 2 Centre for Integrated Genomic Medical Research, University of Manchester, Manchester, UK Correspondence to: Jason J. Brown Centre for Integrated Geno- mic Medical Research University of Manchester Stopford Building, Oxford Road Manchester M13 9PT, UK Tel: þ44 161 275 1622 Fax: þ44 161 275 1617 e-mail: jason.brown@man. ac.uk Abstract: The full extent of the polymorphism of ELA-DRA in Equidae is not yet known. Given the apparent differences in DRA polymorphisms between Equidae and other species, the aims of this study were to more fully characterize ELA-DRA, determine the extent of gene polymorphism and establish the allele-frequency distribution. An allele reference panel for the second exon of ELA-DRA was established by sequence-based typing of 69 equine DNA samples consisting of various breeds of domestic horse (Equus caballus), together with donkeys (Equus asinus), Grant’s zebras (Equus boehmi) and one onager (Equus hemionus). Five of the six previously reported alleles detected using single-strand conformation polymorphism were found: ELA-DRA*0101, ELA-DRA*0201, ELA-DRA*0301, ELA-DRA*0501 (Albright-Fraser DG et al. Polymorphism of DRA among equids. Immunogenetics 1996: 43: 315–7) and ELA-DRA*0601 (GenBank accession number AF5419361). In addition to the previously reported alleles, five novel ELA-DRA alleles were detected within the ELA-DRA allele reference panel. One of these was identified in E. caballus (ELA-DRA*JBH11), one in E. boehmi and E. hemionus (ELA-DRA*JBZ185) and three in E. asinus (ELA-DRA*JBD3, ELA-DRA*JBD17 and ELA-DRA*JBH45). A total of 565 equine DNA samples were screened using reference-strand-mediated conformation analysis, a double-stranded conformation-based mutation detection system that can be used to type existing ELA-DRA alleles and identify new variants. Based on our findings, at least 11 ELA-DRA alleles are now known to exist, and this level of polymorphism at the DRA locus appears to be unique to the genus Equus. Both the previously reported alleles and the new alleles displayed a species-specific distribution. A major function of major histocompatibility complex (MHC) mol- ecules is to bind to antigen peptide fragments and display them on the surface of antigen-presenting cells for recognition by appropriate T cells. In individual species, the degree of polymorphism of each MHC gene locus determines its role in controlling specific immune response. The MHC in all higher vertebrate species is divided into three regions (classes I, II and III), and in those species characterized to date, these regions are located within a tightly linked chromosome region of approximately 4 cM (1). Unlike the MHC in humans (2) and Key words: DRA; ELA; horse; major histocompatibility complex; RSCA Acknowledgments: This work was possible due to the generosity of the Home of Rest for Horses by award of a welfare project grant. The authors thank the Donkey Sanctuary and Stephanie Sanderson MRCVS of the North of England Zoological Society Chester Zoo, for providing blood samples, and Dr Gina Pinchbeck for statistical advice and analysis. J.M. is funded by the International League for the Protection of Horses. Received 28 October 2003, revised 10 March 2004, accepted for publication 29 March 2004 Copyright ß Blackwell Munksgaard 2004 doi: 10.1111/j.1399-0039.2004.00269.x Tissue Antigens 2004: 64: 173–179 Printed in Denmark. All rights reserved 173