奥尼杂交罗非鱼染色体行为研究

Hybridization breeding was traditional breeding technology in aquaculture of our country. It was showed that hybridization and later event(s) played important roles in species evolution by more and more evidence. Hybridization induced chromosome behavior, which was considered a kind of dynamic to evolution. In this study, gynogenesis was induced in O. niloticus and O. aureus using UV-irradiated spermatozoa to activate eggs, then the maternal genome was duplicated with cold shock to prevent it first cleavage. Gynogenesis could select isogenic gynogens immediately. The isogenic hybrids were reproduced by gynogenetic O. niloticus (♀) × O. aureus (♂) at independent net cage. We examined the ploidy, karyotype and sub-chromosome behavior of the isogenic hybrid using flow cytometry, observation of the chromosomes, SSR and HRM, respectively. We want to reveal the general regular patterns of chromosome behavior after hybridization and discuss the relationship between hybridization and evolution. This study could be confirmed some sex-determining region Y in tilapia which had great significance for breeding of tilapia.

杂交育种一直是我国水产养殖业中的传统育种技术。越来越多的证据表明，杂交和杂交后的一系列事件在物种进化过程中也起着非常重要的作用。鱼类杂交后引起的染色体行为变化被认为是进化的动力之一。本项目旨在利用人工诱导雌核发育技术，抑制尼罗罗非鱼和奥利亚罗非鱼第一次卵裂，达到基因座位快速纯化的目的。然后将纯化的尼罗罗非鱼（♀）×奥利亚罗非鱼（♂）一一配对，分别放入不同网箱人工杂交，制备理论上同基因型的奥尼杂交罗非鱼。运用流式细胞术、中期染色体制片技术、微卫星技术及高分辨率融解曲线技术检测奥尼杂交罗非鱼在倍性、核型、亚染色体组水平的染色体行为，揭示杂交后染色体行为的一般规律，探究杂交与物种进化的关系。通过亚染色体组的对比初步确定罗非鱼性别决定相关序列，为进一步的罗非鱼育种打下理论基础。

罗非鱼是世界范围内重要的养殖鱼类，而奥尼杂交罗非鱼是罗非鱼养殖的主要品种。本研究通过紫外灭活精子刺激尼罗罗非鱼和奥利亚罗非鱼人工雌核发育，获得的人工雌核发育罗非鱼经流式细胞术检测确认。将尼罗罗非鱼（♀）×奥利亚罗非鱼（♂）杂交获得奥尼杂交罗非鱼，并自交至F3代，使用组织学切片、流式细胞术和肾细胞染色体制片技术检测F1-F3代奥尼杂交罗非鱼，揭示奥尼杂交罗非鱼为正常的二倍体杂种，行正常两性生殖；对奥尼杂交罗非鱼F1-F3代D-loop序列和GH基因进行了克隆和比对，进一步了解其亚染色体水平的变化规律，发现其线粒体D-loop序列出现重组现象；开展了罗非鱼繁殖生理学研究，克隆了奥利亚罗非鱼vasa基因，并研究了LHRH-A对其表达的影响， 发现LHRH-A对精巢细胞中vasa的表达有抑制作用，揭示LHRH对精巢vasa基因表达有重要调控作用。研究了LHRH-A和hCG对尼罗罗非鱼性腺Piwi表达的影响， 表明Piwis表达的降低可能在精巢分化和发育的过程中起重要作用。本研究揭示了奥尼杂交罗非鱼染色体行为的规律，对罗非鱼育种工作具有重要的意义。