zma-miR159c调控玉米籽粒发育的级联基因表达调控网络研究

Maize kernel contributed most to the biological yield; it was determined during kernel development. In a former work, our group had constructed an over-expression vector of zma-miR159c and successfully transferred it into maize inbred line C01. Phenotyping of T1 and T2 kernels from 15 independent positive transgenic cases showed that kernel length, kernel width an kernel weight of the transgenic lines increased significantly compared to the non-transgenic checks, respectively. The results showed that zma-miR159c should be a key factor influencing maize kernel development and thus an important gene contributed to maize yield. Two genes encoding GAMYB transcription factor (GRMZM2G028054 and GRMZM2G139688) were verified to be the target genes of zma-miR159c via kernel degradome sequencing. In this study, the gene regulating networks of zma-miR159c and the two GAMyb were to be established based on the compare of gene expression data of transgenic and non-transgenic developing kernel samples. The down-stream genes of GAMYB transcription factors were to be detected by combination of ChIP-seq and DAP-seq, followed by the arrangement of these genes into the gene regulating networks. The zma-miR159c dependent zma-miR159c-GAMyb gene expression cascades were then to be established based on the experimental and sequencing data. Our work will contribute both to the theoretical study and technical material of maize breeding.

籽粒是玉米生物学产量最重要的构成因子，而籽粒发育直接决定了籽粒产量。课题组前期利用zma-miR159c过表达载体对玉米自交系C01进行了遗传转化，15个独立转化事件的T1-T2转基因阳性籽粒粒长、粒宽和百粒重等籽粒性状与受体自交系C01相比显著增加，说明zma-miR159c是玉米籽粒发育的调控因子。籽粒降解组测序表明，2个GAMYB转录因子编码基因(GRMZM2G028054和 GRMZM2G139688)是zma-miR159c直接剪切的靶基因。本研究拟构建zma-miR159c及其2个靶基因GAMyb调控玉米籽粒发育的代谢调控网络；筛选GAMYB转录因子调控的下游基因，并阐明这些下游基因在zma-miR159c调控网络中的地位和作用；进而解析zma-miR159c-GAMyb在玉米籽粒发育中的级联基因表达调控网络，为选育优质高产的玉米新品种提供理论与技术支撑。

籽粒是玉米产量最重要的构成因子，而籽粒发育直接决定了玉米产量。本项目以前期研究获得的一个潜在籽粒发育调控因子zma-miR159为研究对象，解析了该上游因子调控玉米籽粒发育的分子路径。通过本项目研究，确证了zma-miR159是玉米籽粒大小和产量的正控制因子；zma-miR159通过控制玉米籽粒胚乳细胞数量而调控玉米籽粒发育。通过遗传验证和分子生物学实验，确定了zma-miR159直接的靶基因为zmMYB74和zmMYB138，且zma-miR159对籽粒细胞数量和产量的调控作用是通过对zmMYB74和zmMYB138的转录后降解作用而实现的。捕获了zmMYB74和zmMYB138直接结合的下游基因启动子序列模体，并验证了4个关键下游基因，确认zmMYB74和zmMYB138通过对下游基因转录激活，控制玉米籽粒发育。总之，本研究解析了“zma-miR159-MYB转录因子-下游基因”级联途径对玉米籽粒大小和产量的调控作用，为选育优质高产的玉米新品种提供了有自主知识产权的基因资源和种质储备，为玉米增产分子育种提供了理论与技术支撑。. 在本项目资助下，发表第一标注的SCI论文2篇，第一标注的中文核心期刊论文3篇，第二标注的SCI论文2篇；获授权国家发明专利2项；培养博士研究生1名，硕士研究生3名；1名项目组成员由中级职称晋升为副高级职称。