扬子鳄环境适应的MHC多样性

Genome-wide single nucleotide polymorphisms (SNPs) of the Chinese alligator (Alligator sinensis) demonstrate that this endangered reptile possesses a nearly identicial level of SNP heterozygosity in intronic regions to that in coding sequences (exons)(Cell Research, 2013, 23: 1091), which indicates that the Chinese alligator has undergone a substantial decrease in the effective population size and has resulted in an extremely low level of genetic diversity. However, the bottlenecked Chinese alligator has not shown declines in its adaptive quality, such as high rates of infection and mortality, making it interesting to determine Alsi-MHC genomic structure and adaptive diversity patterns at different MHC loci. Since the whole-genome sequencing of the Chinese alligator has failed to assemble the Alsi-MHC partial genome, which resulted from a great number of MHC gene copy and high sequence similarities among MHC loci, this project will build a physical map for the Alsi-MHC region based on a ready-to-use BAC (Bacterial Artificial Chromosome) genomic library of the Chinese alligator and identify all of Alis-MHC class I and II loci by BAC clone sequncing. Then, this project will develop multiple suites of locus-specific MHC genotyping techniques and examine adaptive variation patterns of different alligator populations. Finally, this project will explore molecular mechanisms of environmental adaptation in the Chinese alligators from the perspectives of genomic structure and allelic polymorphism.

扬子鳄全基因组层面上的外显子杂合度和内含子杂合度尚处于相同水平，表明物种仍然未摆脱"瓶颈效应"的影响（Cell Research,2013，23:1091）。然长久以来，种群遗传多样性低和近亲繁殖严重的"瓶颈效应"虽已导致尾部畸形问题，但迄今却并未出现高疾病致死率的环境适应性衰退问题。有鉴于全基因组测序技术难以分离确认MHC位点而建立环境适应性探索的功能性分子标记系统，本项目拟在业已完成了扬子鳄基因组BAC文库构建的基础上，进一步通过构建MHC基因组物理图谱及图谱克隆测序、基因注释，MHC基因组组装、位点分离与确认等研究工作，解析MHC的基因组结构，并完整地建立扬子鳄I、II类MHC位点的分子标记系统。继后，通过I、II类MHC分子标记的群体检测分析，自环境适应性免疫功能基因家族的基因组结构和多态性变异等视角，探索扬子鳄适应环境的分子机制这一科学问题。

扬子鳄全基因组层面上的外显子杂合度和内含子杂合度尚处于相同水平，表明物种仍然未摆脱“瓶颈效应”的影响。然长久以来，种群遗传多样性低和近亲繁殖严重的“瓶颈效应”虽已导致尾部畸形问题，但迄今却并未出现高疾病致死率的环境适应性衰退问题。有鉴于全基因组测序技术难以分离确认MHC 位点而建立环境适应性探索的功能性分子标记系统，本项目通过构建MHC 基因组物理图谱及图谱克隆测序、基因注释，MHC 基因组组装、位点分离与确认等研究工作，解析了MHC 的基因组结构，并完整地建立扬子鳄I、II 类MHC 位点的分子标记系统。通过I、II 类MHC 分子标记的群体检测分析，获得如下结果:（1）发现扬子鳄MHC的经典I类区与经典II类区是连在一起，形成2.6M的基因组区段，并从中注释得到101个基因，包含16个I类基因（4个假基因）和17个II类基因（8个假基因），但最终获得PCR产物特异稳定且分型图谱清晰可辨的的3个MHC位点（I1327、I20及Beta1085）。后续的群体分子检测结果表明I1327和I20位点均只有两条等位基因，呈二态，而Beta1085位点有3条等位基因，呈三态。（2）利用研发好的MHC分型技术和本团队已建立的微卫星分型技术对大群体进行分子多态性检测，发现扬子鳄环境适应性的MHC维持机制是平衡选择效应，而目前的样本不足以分析扬子鳄基于MHC的配偶选择机制。