水稻花药发育基因DMT1的克隆与功能分析

Rice was the most important crops in the world, the research on rice anther development will provide the basis for the elucidation of rice reproductive development mechanism and genetic improvement. defective microspore and tapetum1 (dmt1) was a stable genetic male sterile mutant, which was controlled by a single recessive nuclear gene. dmt1 anther was small and white, exhibiting an obvious defect with predegradation tapetum, irregular microspore and abnormal accumulation ROS during the microspore development stage. Using map-based cloning strategy, the DMT1 gene was mapped to a new unreported region-129kb interval on the long arm of Chromosome 1. Base on these findings, a fine mapping will be simultaneously carried out to clone this target gene; the mutation phenotype will be further investigated using cell biology analysis methods; the analysis of the special expression pattern, subcellular localization, transgenic rice and molecular regulator network will be performed to ascertain DMT1 function and molecular mechanism. Through this research, we expected to reveal the relationship between the gene with rice anther development in the theory, to laid foundation and direction for breeding new sterile in application.

水稻是重要的粮食作物，对水稻花药发育的研究将为阐明其生殖发育机制及遗传改良提供基础。defective microspore and tapetum1（dmt1）是前期获得的一个稳定遗传的雄性不育突变体，受单隐性核基因控制。dmt1花药较小呈白色，小孢子发育时期绒毡层提前降解、小孢子形状不规则和活性氧异常累积。通过图位克隆的方法已将其定位于1号染色体长臂上129kb区间内，该区间之前未见花药发育相关基因的报道。本课题拟在已有的工作基础上通过精细定位克隆DMT1基因，细胞生物学方法分析突变表型，基因特定表达模式、亚细胞定位、转基因水稻及分子调控网络的研究阐明其在水稻花药发育中的功能及分子作用机制。通过本研究可望在理论上揭示该基因与水稻花药发育之间的关系，在应用上为选育出新的不育系奠定基础和指明方向。

水稻(Oryza sativa L.)是人类最主要的粮食作物之一。雄性不育是雄性器官发育异常或退化，不能形成有活力的花粉而雌蕊发育正常的现象。雄性不育在水稻杂交育种和杂种优势利用中处于核心地位，在保障国家粮食安全上意义重大，对水稻雄性不育的相关机制的研究具有一定的理论和实践意义。dmt1(defective microsporocyte tapetum1) 是一个通过60Coγ射线辐射诱变松香早粳获得的一个稳定遗传的雄性不育突变体。细胞生物学的分析表明，dmt1突变体在花药发育过程的主要缺陷是绒毡层提前降解、乌氏体和小孢子外壁不能正常行程，导致花粉败育。通过图位克隆结合重测序的方法，最终将DMT1基因定位在19kb的区间内，该区间内仅含一个包含DUF2215结构域的功能未知蛋白。通过序列测定发现dmt1突变体中目的基因第5外显子/内含子剪切处由G变为A，由于转录本的异常导致蛋白提前终止。DMT1在花药发育的各个时期都能检测到表达，并在小孢子时期达到最大值，原位杂交分析表明其主要在绒毡层和小孢子中表达。转录本的分析表明野生型和dmt1突变体间差异性表达基因主要富集于核糖体、角质蜡质相关代谢及苯丙烷合成途径，推测DMT1可能还参与花粉外壁合成。通过本研究可加深对水稻花药发育分子机制和调控网络的理解，在应用上为水稻的遗传改良提供理论依据。