ANALYSIS OF ZEBRAFISH MHC USING BAC CLONES

Although major histocompatibility complex (Mhc) genes have been identi fied in a number of species, little is yet known about their organizat ion in species other than human and mouse. The zebrafish, Danio rerio, is a good candidate for full elucidation of the organization of its M hc. As a step toward achieving this goal, a commercially available zeb rafish BAC library was screened with probes specific for previously id entified zebrafish class I and class LI genes, as well as for genes co ntrolling the proteasome subunits LMP7 and LMP2. Restriction maps of t he individual positive clones were prepared and the Mhc (LMP7) genes l ocalized to specific fragments. The total length of genomic DNA fragme nts with Mhc genes was approximately 1700 kilobases (kb) (200 kb of fr agments bearing class I loci and 1500 kb of fragments bearing class II loci). One of the two class I loci (Dare-UCA) is closely associated w ith the LMP7 locus: the second class I locus (Dare-UAA) is more than 5 0 kb distant from the UCA locus and has no LMP genes associated with i t. None of the class II genes are linked to the class I or the LMP gen es. All six of the previously identified class II B genes and one of t he three class II A genes were found to be present in the BAC clones; no new Mhc loci could be identified in the library, Each of the six pr eviously identified class II B loci was found to be borne by a separat e group of BAC clones. The Dare-DAB and -DAA loci were found on the sa me clone, approximately 15 kb apart from each other. An expansion of D CB and DDB loci was detected: the zebrafish genome may contain at leas t five closely related DCB and two closely related DDB loci which are presumably the products of relatively recent tandem duplication. These results are consistent with linkage studies and indicate that in the zebrafish, the class I and class II loci are on different chromosomes, and the class II loci are in three different regions, at least two of which are on different chromosomes.