一个水稻花粉不育新基因的图位克隆与功能分析

Pollen sterility that affects seed setting rate of spikelet is crucial for rice yield . Male-sterile mutations are ideal materials for studying the molecular mechanism of pollen development in rice. Research on male-sterile mutants provides a theoretical basis for the application of male-sterile materials in breeding and production. In earlier stage of this research, we found a rice male-sterile mutant. Pollen fertility and seed setting rate of the mutant were about 9% and 5%, respectively, that were significant differences of the wild-type. Genetic analysis indicated that monofactorial recessive gene controlled male-sterile inheritance in the mutant. By extreme individual mapping method, we have restricted the candidate gene to about 88-kb between markers InZ-3 and InZ-4 on rice chromosome 3. Gene predicting indicated no allele conflicts with any rice male-sterile genes reported. Therefore, we believe that it is a novel gene conferring rice pollen sterility. By a sequence analysis of these predicted genes, we found only a single nucleotide convert in the coding region of a gene, which caused change of a conserved amino acid. So we presumed this gene should be the candidate. The project intends to separate the male-sterile mutant gene via map-based cloning method and study the function of the male-sterile gene to clarify the molecular mechanism of how it regulates rice pollen sterility.

花粉育性直接影响小穗结实率，是影响水稻产量的关键因素之一。花粉不育突变体是研究水稻花粉发育分子机制的理想材料，水稻花粉不育突变体的研究可为花粉不育材料在育种和生产上的应用提供理论依据。本项目前期发现一个水稻花粉不育突变体，该突变体的花粉育性、小穗结实率分别为9%、5%，较野生型有显著差异。遗传分析表明，该突变性状受隐性单基因控制。通过极端个体定位方法，已将该基因精细定位在水稻第3染色体标记InZ-3和InZ-4之间约 88-kb的区间内。基因预测表明，定位区间内基因与已报道的水稻花粉不育基因均不等位，该基因是一个新的水稻花粉不育基因。通过对预测基因进行测序分析，发现有一个基因在突变体中发生了一个碱基的替换，导致一个保守氨基酸的改变，将该基因确定候选基因。本项目拟通过图位克隆的方法分离该突变基因，并对其功能进行较为系统的研究，探索其对水稻花粉不育的分子调控机理。

水稻雄性不育普遍存在，严重影响农业生产。花粉发育相关基因的突变可以导致花粉小孢子的发育异常，最终不能形成成熟花粉粒，从而影响小穗结实率。分离和克隆花粉发育基因有助于我们进一步了解花粉发育的分子机理。本研究以花粉不育突变体ps3为材料，表现为花粉育性显著降低，小穗结实率极低。通过细胞学观察，我们发现突变体ps3的花药壁能够正常形成和及时降解，突变体ps3的小孢子发育在游离小孢子时期出现了异常，导致小孢子发育停滞在单核期。利用遗传互补实验，我们证实PS3基因就是控制花粉发育表型的目的基因。PS3基因在水稻各器官中呈组成性表达，在单核期的花药中表达量最高。PS3蛋白被定位在细胞核和细胞质中。在突变体ps3的成熟花药中，GBSS1基因的表达量显著下调，说明PS3基因可能与淀粉合成有关。总之，以上结果让我们对PS3基因调控水稻花粉育性和小穗结实率有了新的认识，该基因在小孢子发育过程中具有非常重要的作用。