VH REGION DIVERSIFICATION AMONG HAPLOTYPES

描述：(改编自调查人员摘要)长期目标是 要了解VH地区多样化背后的机制 人免疫球蛋白重链基因复合体及其作用 多样化对人类健康的影响。工作假设是，这些差异 在这一地区的多样性所产生的具有重大生物学意义的 和遗传暗示，并可能包含遗传基因留下的“足迹” 导致多元化的事件。对这些足迹的研究可能会有所帮助 了解造成VH地区多样化的机制。这个 VH地区是高度多元化的。人们认为没有单倍型包含 一组相同的VH基因片段。因此，即使一个完整的核苷酸序列 因为VH区是通过使用二倍体材料产生的，所以序列可能不 代表任何单倍型的实际序列，远远不足以 了解VH区域的物理结构。在拟议的研究中，50 具有高度多样化遗传背景的男性人类受试者将是 被招募了。供者单倍型中VH基因片段的数目和组成 将通过分析来自这些捐赠者的单个单倍体精子细胞来确定。 将根据选举结果选出四个有代表性的捐助者。单倍型 将确定这四个受试者中VH基因片段的组织 通过分析单个精子的单个DNA片段。由此产生的信息 将用于组织来自噬菌体的含有VH基因片段的克隆 这些捐赠者的基因组文库。所有VH基因片段和 它们的侧翼区域包含启动子、转录调控元件、 并确定每个单倍型中的重组信号序列。这个 将检查每个单倍型的详细物理结构，以确定 限制性图谱绘制后的遗传兴趣区。序列中的 将确定已确定的地区。生成的数据将用于 推断其余46例单倍型中的VH基因组织 个人和进一步分析VH地区多样化的影响 以及多元化背后的机制。中开发的方法 该研究也可用于分析其他类型的单倍型基因组织 多基因家族，包括那些参与免疫发展的基因 系统。并确定染色体的详细物理结构 具有单拷贝序列的区域。

DESCRIPTION: (Adapted from investigator's abstract) The long-term objective is to understand the mechanisms underlying diversification of the VH region in the human immunoglobulin heavy chain gene complex and the effect of the diversification on human health. The working hypothesis is that the differences resulted from the diversification in this region are of significant biological and genetic implications, and may contain "footprints" left by the genetic events responsible for the diversification. Study of these footprints may help to understand the mechanisms responsible for the VH region diversification. The VH region is highly diversified. It is believed that no haplotypes contain the same set of VH gene segments. Therefore, even if a complete nucleotide sequence for the VH region is generated by using diploid materials, the sequence may not represent the actual sequences in any haplotypes and is far from sufficient for understanding physical structure of the VH region. In the proposed study, 50 male human subjects with highly diversified genetic backgrounds will be recruited. VH gene segment number and composition in the donors' haplotypes will be determined by analyzing single haploid sperm cells from these donors. Four representative donors will be selected based on the results. The haplotype organization of the VH gene segments in these four subjects will be determined by analyzing single DNA fragments from single sperm. The resulting information will be used to organize the VH gene segment-containing clones from phage genomic libraries for these donors. The sequences of all VH gene segments and their flanking regions containing promoters, transcription regulatory elements, and recombination signal sequences in each haplotype will be determined. The detailed physical structure of each haplotype will be examined to identify the regions of genetic interest after restriction mapping. The sequences of the identified regions will be determined. The resulting data will be used to deduce the VH gene organization in the haplotypes of the remaining 46 individuals and for further analysis impact of the VH region diversification and the mechanisms underlying the diversification. The approaches developed in the study may also be used to analyze haplotype gene organization of other multi-gene families including those involved in the development of the immune system. And to determine the detailed physical structure of the chromosomal regions with single copy sequences.