Localization of bovine lymphocyte antigen (BoLA) DYA and class I loci to different regions of Chromosome 23

Abstract

The class II loci of the major histocompatibility complex (MHC)of humans (HLA) and mice (H-2) are embedded among other lociof the MHC, which spans approximately 4 mb of DNA and has arecombination length of about 2 cM (Campbell and Trowsdale1993; Himmelbauer et al. 1993).In contrast, recombination analysis of class II genes in thebovine lymphocyte antigen complex (BoLA) revealed two distinctclusters of class II loci about 17 cM apart (Anderson et al. 1988).One cluster designated IIa contains the bovine homologs of thehuman DQ and DR gene families and is tightly linked to otherBoLA genes, including HSP70, CYP21, and several class I loci(Bishop et al. 1994). The second cluster, IIb, contains the DYA,DYB, DOB loci and a novel class II gene designated DIB (Stoneand Muggli-Cockett 1990). The disruption of the BoLA class IIloci into two clusters suggests that the BoLA complex has under-gone significant rearrangements not yet observed in the MHCs ofhumans or mice, but that apparently have occurred in the MHCs ofchickens (Miller et al. 1994) and swine (Smith et al. 1995). Alter-natively, the bovine class IIa and IIb gene clusters could be inphysical proximity but undergo frequent recombinant owing to oneor more recombination hotspots. Recombination hotspots havebeen reported in the HLA and H-2 regions (Shiroishi et al. 1990;Steinmetz et al. 1986; Yoshino et al. 1994), and anomalous re-combination frequencies in this region among individual bullshave been reported (Park et al. 1995).To test whether the BoLA IIb cluster mapped to a differentchromosomal location than the other BoLA loci, we performedfluorescence in situ hybridization (FISH) analysis of bovine Chro-mosome (Chr) 23, using large insert clones containing DYA orclass I sequences as fluorescent probes. In this report, we demon-strate that DNA probes containing bovine DYA sequences map toa different region of Chr 23 than does DNA containing class Isequences, consistent with the reported recombination distance of17 cM between these genes.Recombinant clones of bacterial artificial chromosomes(BACs) containing DYA or class I sequences were isolated byPCR screening of a bovine BAC library (Cai et al. 1995) withprimer pairs that have been previously described (Van Eijk et al.1992; Garber et al. 1994). Authenticity of each clone was con-firmed by sequencing of the respective PCR products and com-paring with published sequences.BAC clones 6 and 76 were identified by PCR as containingDYA and class I sequences, respectively, and DNAs from theseclones were prepared for FISH. The sequence of the PCR productfrom BAC 6 differed at only a single position,