Information on the relative order of Igk-V gene families has come from studies in inbred mice and their recombinants of V kappa phenotypic markers and restriction fragment length polymorphisms (RFLPs) associated with individual gene families. Information on the nature of the polymorphism has come from cloning, restriction mapping and nucleotide sequence analysis of allelic members of gene families from inbred and wild mice. Such studies provide information on the evolution and dynamics of the Igk-V locus which has relevance to loci encoding immunoglobulin VH regions, T cell receptor V regions, and major histocompatibility complex genes of mouse and man. Previous studies from this laboratory have characterized four alleles at the Igk-VSer locus in Mus. m. domesticus which differ in the genes themselves and in the distribution of repetitive and unique DNA upstream of the genes.
Specific aims of the proposed studies are: 1) to clone and compare the structures of new Igk- VSer alleles identified in wild mice; 2) to use phage and cosmid approaches to """"""""walk"""""""" upstream and downstream of Igk-VSer alleles to further investigate the basis for their divergence; 3) to clone and analyze additional genes detected with a larger V kappa Ser cDNA probe to determine whether they belong to the Igk-VSer gene family; 4) to use field inversion gel electrophoresis (FIGE) and contour-clamped electric field electrophoresis (CHEF) together with Southern hybridization to establish the long range order of genes within and closely linked to the Igk-V locus; 5) to analyze the polymorphic genes of the V kappa 10 group and determine why V kappa 10 alleles linked to the Igk-VSer alpha allele cannot contribute to expression of the major cross-reactive anti-p-azophenylarsonate idiotype of A/J mice; 6) to continue the production and analysis of new BALB/c-congenic strains which bear newly-identified Igk-VSer alleles and linked chromosome markers; 7) to determine the nucleotide sequence of new Igk-J alleles identified in wild mice; and 8) to produce hybridomas with spleen cells of mice bearing different Igk-VSer alleles, to identify ones producing V kappa Ser and V kappa 10 light chains by dot hybridization to RNA and to determine their nucleotide sequence diversity for comparison with the germ line alleles.
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