鸡TCRβ和TCRγ基因位点结构和组库多样性分析

项目名称： 鸡TCRβ和TCRγ基因位点结构和组库多样性分析

项目编号： No.31501943

项目类型： 青年科学基金项目

立项/批准年度： 2016

项目学科： 畜牧学与草地科学

项目作者： 孙亿

作者单位： 山东农业大学

项目金额： 20万元

中文摘要： T细胞受体（TCR）作为T淋巴细胞识别内/外源性抗原的关键分子，在免疫应答和免疫调节中发挥着重要的作用。迄今为止，对TCR基因结构特征及组库多样性的认识基本上是以人类自身或者小鼠作为实验模型获得的，对于这些分子机制在家养动物，特别是家禽中的认识还很少。本研究拟通过鸡基因组BAC文库筛选和阳性BAC克隆全长测序的方法，获得鸡TCRβ和TCRγ基因位点的完整结构，并对其中恒定区（C）和可变区（V）基因片段进行结构特征及遗传进化关系分析。同时，利用流式细胞分选获得7-8周龄SPF鸡外周血、脾脏和盲肠上皮内的CD4+CD8-αβ、CD4-CD8+αβ、CD8-γδ和CD8+γδT细胞亚群，应用5’RACE和Illumina Hiseq 2500高通量测序技术获得各T细胞亚群表达的TCR组库，分析各T细胞亚群TCR多样性特点，为阐明家禽TCR的进化规律及多样性产生的分子机制提供理论依据。

中文关键词： 鸡；T细胞受体；T细胞亚群；组库；多样性

英文摘要： T cells, which play a major role in regulation and effector functions of immune responses, have T-cell receptor (TCR) heterodimers expressed on the surface to react with either antigenic peptides presented by major histocompatibility complex molecule (for αβTCR) or a variety of antigens irrespective of their molecular or genetic nature (for γδTCR). So far, the genomic structure and repertoire diversity of the TCR genes have been extensively studied in both humans and mice but rarely in domestic animals, especially in poultry. In this study, we will first obtain the complete genomic structure of the chicken TCRβ and TCRγ loci using shotgun sequencing of BAC clones, and then analyze the sequence characteristic and phylogenesis of V and C segments. Subsequently, the CD4+CD8-αβ、CD4-CD8+αβ、CD8-γδ, and CD8+γδ T cell subsets will be sorted by FACS from peripheral blood, spleen and cecal epithelia from SPF chicken with 7 to 8 weeks old. The TCR repertoire from individual T cell subset will be evaluated using 5’RACE assay and Illumina Hiseq 2500 high-throughput sequencing, and the repertoire diversity of individual T cell subsets will be analyzed and compared. This study will provide new understanding for the evolution and diversity of poultry TCR genes.

英文关键词： chicken;T cell receptor;T cell subset;repertoire;diversity