油松球花发育的分子调控网络构建及关键基因筛选

Up to now, the molecular regulatory network on the flower development of angiosperm plant species has been established. However, in gymnosperms species, studies on the development of reproductive organs are quite inadequate compared with angiosperms due to the limitations caused by long time in growth and development, the huge genomes and limited genome information in the Genebank. All the existing research works about molecular mechanism of reproductive organs development of gymnosperms were only tracking with the progress on angiosperms. That leads to the results that molecular mechanism of reproductive organs development and the key genes are largely unknown and can not provid the basic theory for genetics improvement and strobile development in conifers. In this project, microarrays will be designed based on the transcriptome database of P.tabuliformis which is a native gymnosperm tree species in China. Through microarray analysis of large scale gene expression profiles between male and female cones at the different stages of development and miRNA and RNA degradome sequencing analysis of both bisexual and normal cones, the differences and patterns of gene expression during the dynamic developmental process in male and female cones will be characterized, the co-expression and regulation network database of genes associated with the development of male and female cones will be built, and the key genes involved in the regulation of male and female cones development and the mutation of bisexual strobilus will be identified. These studies will provide theory basis for genetic improvement of P.tabuliformis and also make new experimental evidence and molecular basis for evolutionary mechanism of reproductive structures in higher plants of gymnosperms and angiosperms.

被子植物生殖器官发育的分子调控网络已经建立，裸子植物由于生长周期长、基因组大、可利用基因组信息少，其生殖器官发育的分子基础研究只能跟踪被子植物的研究成果，导致了裸子植物生殖器官发育的基因表达调控不清楚，关键基因不能确定，对针叶树种的遗传改良和雌雄球花发育难以起到有效的理论支撑。本课题以裸子植物油松为研究对象，在已有转录组数据库的基础上，制备油松表达谱芯片，以油松正常球花和两性球花发育过程关键时期获取的材料为基础，通过油松表达谱芯片杂交，结合miRNA与降解组测序及生物信息学分析方法，开展油松雌雄球花发育动态过程中基因差异表达规律研究，构建雌雄球花发育相关基因的共表达及调控网络数据库，确定油松雌雄球花形成的关键基因。研究结果不仅为油松的遗传改良提供理论基础，也为种子植物生殖器官的分子进化研究提供理论基础和基因信息。

生殖器官从发育早期到成熟的不同阶段蕴含了非常丰富的分类、系统和进化发育信息，是高等植物发育和进化研究的核心。裸子植物由于生长周期长、基因组大、可利用基因组信息少，其生殖器官发育的分子基础研究只能跟踪被子植物的研究成果，导致了裸子植物生殖器官发育的基因表达调控不清楚，关键基因不能确定，对针叶树种的遗传改良和雌雄球花发育难以起到有效的理论支撑。本研究以裸子植物油松为研究对象，利用Roche 454与Solexa两种测序平台分别对多种油松组织与多个发育阶段雌雄球花样本进行了深度测序，构建了油松高质量参考转录组序列；明确了雄球花发育调控在油松两性分化中的核心作用，并确定了MADS-box基因是其中的核心调控因子，构建了油松球花发育的分子调控网络；发现油松中至少存在23个II型MADS-box基因，其中包括4个B类与1个C类功能基因，并且首次从针叶树中分离雌球花特异表达的MADS-box基因；证实miRNA是油松主要的sRNA，发现参与被子植物生殖发育调控的多个miRNA-靶基因调控通路在油松中同样是保守的，获得26个确定存在相互作用的miRNA家族与对应的74个靶基因，克隆获得了其中36个靶基因的全长CDS序列；揭示了针叶树两性球花的雄性来源，为两性进化起源“OOM”假说提供了实验证据，同时也证实油松球花性别调控雄球花原基遗传操作策略的可行性。研究结果不仅为油松的遗传改良提供理论基础，也为种子植物生殖器官的分子进化研究提供理论基础和基因信息。