The goal of this study is to identify the functional diversity (D) segments of the human immunoglobulin heavy chain (IgH) gene and to obtain insights into the molecular mechanism of VDJ recombination in this gene. Experiments are proposed herein to study the entire repertoire of human functional D segments by molecular cloning of functionally rearranged IgH sequences and to characterize the genomic organization of the whole repertoire of D segments. An in vivo assay system of DJ and VDJ recombination will be developed and used to elucidate the molecular basis of VDJ recombination. For this purpose, a series of DNA constructs will be made including those with mutated 7mer-9mer signal sequences and spacer sequences and recombination activity will be assayed in the in vivo system. Analyses are also proposed to determine the underlying basis of V segment joining to the DJ complex, by introducing a DNA construct into the assay system.
