澳洲野生稻EE染色体组特异标记的鉴定及其在稻属种间杂种研究中的应用

In the genus Oryza, O.australiensis originated from northern Australia has the biggest genome in the diploid wild species, and is an unique wild specie with EE genome. As a good monocot model system, O.sativa with AA genome has been extensively studied for decades in molecular biology and cytogenetics, but related research in O.australiensis with EE genome is few. It owes to the weak foundation of molecular biology and cytogenetics, which hinders the mining and utilization of the useful genes of O.australiensis. Using the embryo rescue following hybridization, we have produced interspecific hybrids (F1) and its backcross progeny between O.sativa and O.australiensis. In this research program, we will develop a high-density molecular markers for EE genome and a set of cytogenetics markers for different chromosome arms of EE genome. Combined with the backcross and embryo rescue, introgression lines carrying wild rice (O.australiensis) segments in cultivated rice (O.sativa ) background will be constructed and identified. The functional centromere sequences of O. australiensis will be isolated and identified by means of screening the subclone library and ChIP-Seq using the anti-CENH3 antibody. The difference of gene expression in the meiotic of the aboved genetic materials will be analyzed by means of RNA-seq and qPCR. These results will lay the foundation for fast recognition and tracking of the wild rice (O.australiensis) segments, and it will promote the transfer and utilization of favored genes from O.australiensis to O.sativa.

澳洲野生稻拥有目前稻属二倍体种中最大的染色体组，也是稻属唯一具有EE染色体组的稻种。虽然AA染色体组亚洲栽培稻的分子生物学研究相当深入，但是EE染色体组澳洲野生稻的相关研究还很少，这主要是澳洲野生稻的基础研究还十分薄弱，进而限制了人们对澳洲野生稻有利基因的挖掘和利用。在前期已获得亚洲栽培稻和澳洲野生稻的种间杂种及其后代的材料基础上，本研究拟发展一套高密度覆盖EE染色体组的PCR标记体系；发展一套识别EE染色体组不同染色体（臂）的分子细胞学标记；结合亚克隆和ChIP-seq技术，解析澳洲野生稻功能性着丝粒的序列组成和结构特征；利用回交和胚拯救技术，构建一批以亚洲栽培稻为背景的澳洲野生稻染色体片段渐渗系；借助RNA-seq和定量PCR技术，分析杂种与亲本在穗分化不同时期基因表达模式的差异。研究结果将为快速识别和鉴定澳洲野生稻染色体片段和加快澳洲野生稻有利基因的转移和利用奠定坚实的基础。

作为稻属资源中拥有EE染色体组的稻种，澳洲野生稻主要分布于澳洲北部，其基因组是目前稻属二倍体种中最大的，当前有关澳洲野生稻的研究较少。利用远缘杂交和胚拯救技术，我们前期已经获得了亚洲栽培稻和澳洲野生稻的种间杂种及其后代。在此基础上，本项目发展了一套包含231个高密度覆盖EE染色体组的PCR标记体系；发展了一套识别EE染色体组不同染色体（臂）的分子细胞学标记；结合亚克隆和ChIP-seq技术，鉴定了CentO是澳洲野生稻功能性着丝粒重要元件；利用回交和胚拯救技术，构建一批包含41个以亚洲栽培稻为背景的澳洲野生稻染色体片段渐渗系；借助幼穗RNA-seq技术，发现有2142个基因在杂种与亲本幼穗分化早期差异表达，其中290个基因上调表达，1852个基因下调表达。研究结果将为快速识别和鉴定澳洲野生稻染色体片段和加快澳洲野生稻有利基因的转移和利用奠定坚实的基础。