水稻耐热相关microRNA的鉴定及功能研究

High temperature has become one of the major disastrous climate in influencing crop yields. microRNA (miRNA) has been known to play pivotal roles in gene regulation networks of many important traits of crops. However, no results have been reported for the role of miRNAs in rice heat tolerance. In our former study, from 28 rice varieties of 11 countries, we have screened out a strong heat tolerance variety "Gan-xiang-nuo" during heading stages. Moreover, We have constructed a library of single segment substitution lines (SSSLs) in rice. The library was constructed by using "Hua-jing-xian 74" as genetic background. Based on these studies, this project aims to carry out the following researches: (1) Using strong heat tolerance variety "Gan-xiang-nuo" and heat sensitive variety "Hua-jing-xian 74" as our research material, we expect to screen for differential expressed miRNA to heat stress by applying Solexa sequence technique on the standard of whole genome; then, we will verify the heat tolerance related candidate miRNA molecules combined with the whole genome SSSL group. (2) Function analysis for heat tolerance related candidate miRNA. The implementation and completion of this project not only improve our understanding of the interaction between miRNA and environmental stresses, but also can be used as molecular marker application to molecular breeding in rice heat tolerance.

高温热害已成为影响农作物生产的主要灾害性气候之一。microRNA（miRNA）在作物许多重要性状相关的功能基因调控网络中起着非常重要的作用，但有关miRNA在调控水稻耐热方面还未见有报道。我们前期从全球11个国家的28份水稻资源中筛选出了1份抽穗开花期耐热性很强的籼稻品种"赣香糯"，并且获得了以不耐热性籼稻品种"华粳籼74"为遗传背景的单片段代换系（SSSL）群体。在此基础上，本项目拟开展以下研究：（1）以强耐热品种"赣香糯"和不耐热品种"华粳籼74"为研究材料，通过从全基因组水平测序筛选热害胁迫差异表达的miRNA分子，并结合全基因组SSSL群体鉴定耐热相关候选miRNA；（2）候选miRNA的耐热功能研究。该项目的实施和完成不仅可以提高人们对于miRNA与环境胁迫互作的认知，而且可作为分子标记应用于水稻耐热性分子育种

高温热害已成为影响农作物生产的主要灾害性气候之一。microRNA（miRNA）在作物许多重要性状相关的功能基因调控网络中起着非常重要的作用，但有关miRNA在调控水稻耐热方面还未见有报道。我们前期从全球11个国家的28份水稻资源中筛选出了1份抽穗开花期耐热性很强的籼稻品种“赣香糯”，并且获得了以“赣香糯”为供体亲本，以不耐热性籼稻品种“华粳籼74”为遗传背景的单片段代换系（SSSL）群体。通过对这两个供体亲本以及SSSL群体进行耐热性鉴定，获得了与穗期耐热相关的4个QTL座位。同时，我们通过对强耐热水稻品种“赣香糯”以及不耐热水稻品种“华粳籼74”热处理前后的小穗进行小RNA大规模测序分析，鉴定到109个与水稻抽穗期热响应相关的microRNA。此外，我们还发现，与热敏感品种“华粳籼74”相比，强耐热品种“赣香糯”中热响应microRNA的数目明显增多，并且microRNA对高温胁迫响应的速度也明显快于“华粳籼74”。这些结果表明microRNA在水稻耐热中发挥了非常重要的作用。随后，我们用热响应microRNA与前面鉴定到的耐热相关的4个QTL作图相结合的方法，筛选到了26个候选microRNA，它们有8个靶基因位于QTL区间内。我们挑选了其中的1个microRNA（miR169r-5p）进行转基因功能验证。已有的结果显示与野生型植株相比，miR169r-5p增量表达株的穗期耐热性明显增强，表明miR169r-5p在水稻穗期耐热中发挥正向调控作用。本研究是国内外首次开展水稻抽穗开花期耐热相关miRNA的鉴定的研究，研究结果对深入了解水稻耐热的分子机制和分子育种具有重要意义。本项目把RNA测序与QTL作图结合鉴定水稻耐热相关miRNA及其靶基因的鉴定，提高了效率和准确性，为同类研究提供了一种可以借鉴的新方法。