The goal of this research is to determine the genetic basis for the allotypes of the variable region of rabbit immunoglobin heavy chains. Rabbit B cell leukemias will be developed in transgenic rabbits carrying the c-myc-heavy chain enhancer gene construct. Cell lines will be developed from these rabbits and a somatic cell fusion partner will be developed for the generation of rabbit hybridomas. Rearranged VDJ genes will be cloned from the leukemic cells as well as various hybridomas in order to determine which germline genes are used in VDJ rearrangements and if there is preferential usage of one or a small number of germline V genes. These studies should provide insight into the genetic basis for the apparent allelic inheritance of rabbit VH allotypes. The genetic basis for latent VH allotypes will also be examined by preparation of a cDNA library from rabbits expressing large quantities of a latent a2 allotype. The nucleotide sequence of the cDNAs will be determined and the cDNAs will be expressed in vitro to establish which ones encode the latent a2 allotype. If a latent cDNA is cloned, the corresponding germline V gene will be cloned and analyzed by nucleotide sequence analysis and for its location within the VH chromosomal region. Finally, VHa+, and Vha- cDNA will be cloned and the nucleotide sequence determined in order to understand the structural correlates of VH allotypes. The regulatory region of genes encoding nominal VHa+, VHa- and latent allotypes will be compared in an effort to understand how the expression of the multiple VH genes is regulated.
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