High-Resolution Characterization of the Canine DLA-DRBI Locus Using Reference Strand-Mediated Conformational Analysis.

Kennedy, L. J.; Quarmby, S.; Fretwell, N.; Martin, A. J.; Jones, P. G.; Jones, C. A.; Ollier, W. E. R.; Murphy, William

Several methods exist for genotyping class II DLA gene polymorphisms in the dog. The most accurate method is sequence-based typing, which involves direct sequencing of polymerase chain reaction products. However, this method is expensive and unsuitable for large scale studies. Recently, reference strand mediated conformation analysis (RSCA) has been shown to be effective for characterizing major histocompatibility complex genes in humans, sheep, horse, and cats. RSCA is a cheap and rapid method, ideal for large epidemiological studies. We have developed RSCA for typing DLA DRB1 in the dog. Control panels including dogs typed by sequence based typing and cloned major histocompatibility complex class II alleles in plasmids were used to establish migration patterns for each allele using 20 different fluorescent labeled references, of which 5 were selected to allow for clear identification and discrimination of all known DLA DRB1 alleles. We have compared 168 dogs typed by RSCA for DLA-DRB1 and characterized by sequence-based typing, with less than 1% discrepancy. These differences were due to missing alleles because of a weak polymerase chain reaction. To date, we have RSCA-typed 1,394 dogs. RSCA is likely to become the method of choice for characterizing DLA genes in the dog and will prove a useful tool for dissecting the immune response of dogs in clinical studies.

DOGS; GENETICS; PHENOTYPES; IMMUNOGENETICS; GENETIC polymorphisms; DNA polymerases; POLYMERASE chain reaction; POLYMERIZATION; HLA histocompatibility antigens

Journal of Heredity, 2005, Vol 96, Issue 7, p836

0022-1503

Academic Journal

10.1093/jhered/esi112