水稻spr1突变体提高重组频率的分子机制研究

In the previous study, we have identified MSH5, a meiotic homologous recombination element, in rice. In msh5, the nondisjunction of homologous chromosomes in meiocytes were observed due to the reduction of chiasmata between homologues chromosomes, leading to the serious male and female gamete abortion. Furthermore, we identified the msh5 suppressor, named spr1. In the msh5/spr1 double mutant, almost no univalent occurred and the fertility of msh5 well restored. In this proposal, we are going to perform detail investigation of chromosome behaviors and immunodetection of marker proteins on meiotic chromosomes in both spr1 single and double mutants. We also intend to screen the proteins that interact with SPR1 using Yeast Two-Hybrid and Co-immunoprecipitation strategies, and investigate the increase of genetic recombination frequency in spr1 background using mapping populations. By these studies, we seek to reveal their coordinated mechanisms on the regulation of homologous recombination in rice meiosis. Moreover, we may also get some knowledge on increasing the efficiency of rice breeding by manipulation of SPR1.

MSH5是我们实验室克隆的一个影响减数分裂同源重组的基因，由于该突变体重组频率显著减少，形成大量单价体，最终导致同源染色体不均等分离而产生败育配子。我们在实验室多年从事水稻减数分裂研究的基础上，克隆了可有效提高msh5突变体重组频率基因spr1。研究表明msh5/spr1双突变体同源染色体配对与重组正常，育性正常。本课题拟在此基础之上，通过细胞学和遗传学方法，详细研究spr1突变体提高重组频率的效率；制备SPR1蛋白抗体，通过免疫荧光染色反应研究SPR1在减数分裂染色体上的定位；结合酵母双杂交和免疫共沉淀等技术，深入研究SPR1参与水稻同源染色体重组的分子机理；并在spr1背景下构建分离群体，探索在水稻杂交育种过程中，利用spr1提高杂交后代重组频率的可能性。相关研究结果不仅增加对减数分裂遗传调控分子机理的理解和认识，更为通过改良重组频率提高育种效率提供对策。

重组频率的可控性一直是育种学家们孜孜以求的目标，尤其是提高重组频率有利于打破不良基因的遗传累赘，加速育种进程。由于缺乏明显的形态学筛选标准，采用正向遗传学的方法来筛选重组频率提高、又不影响育性的突变体显然是行不通的。我们通过筛选msh5的抑制子克隆了可有效提高msh5突变体重组频率基因SPR1。在spr1突变体中，减数分裂染色体行为同野生型没有明显差异，而且标识减数分裂早期重组情况的指示蛋白也均能正常定位在染色体上。spr1可以恢复包括msh5在内的所有zmm突变体二价体减少的表型。与野生型相比，spr1大约可以提高1.5倍的重组频率，而且这些增加的交叉结（Crossover）不能被一类交叉结指示蛋白HEI10所识别，表明spr1可以提高二类交叉结的水平。spr1提高重组频率依赖于PAIR1、COM1和DMC1蛋白功能的协同作用。