水稻光合速率主效QTL的精细定位与基因发掘

It has been a hotspot problem all through the world to study the hereditary mechanism of high photosynthetic efficiency genes and breeding of high photosynthetic efficiency in rice,because it is helpful to resolve the problem of food crisis and safety.We had studied systematically the photosynthetic traits of rice and located preliminary the main effect QTLs of rice photosynthetic rate（PR）;Then we breeded the near-isogenic lines( NILs) of main effect QTLs of rice photosynthetic rate by MAS and backcross(158 lines of BC3F2), and constructed the secondary separation populations by crossing the Chromosome Segment Substitution Lines( CSSLs) corresponding to the objective QTLs and the background parent(have gotten 37 objective lines of BC1F1); Then we plan to fine mapping the main effect QTLs of rice photosynthetic rate by aids of using all kinds of modern molecular biological technology such as using high flux RNA-Sequence to get the loci of SNP and the genes which were quickly respond to the light intensity,or utilizing whole genome sequence(solexa technology) to acquire the new markers and forecasting genes which were obtained by analyzing of the sequence of objective chromosome segment and the difference between the parents in the objective chromosome segments, and designing or developing new marks by selecting the SNP loci in the objective chromosome segments, then measure the PR of F2 poulations in two seasons; The next step we will determine the candidate genes of PR by using the analysis of bioinformatics ,analyzing technology of comparative genome and the results of the main effect QTLs of PR in the F2 population; At last, we will carry out the expressing analyse and preliminary functional analysis to the candidate genes,and so we will discover the high photosynthetic efficiency genes in the rice to establish the base of studying on the genetic mechanlism of high photosynthetic efficiency genes of rice and its breeding of MAS.

水稻高光效基因遗传机理及高光效分子育种一直是国内外研究的热点问题，有助于粮食危机问题的解决。多年来我们对水稻光合生理性状进行了系统的研究，对水稻光合速率（PR）主效QTL进行了初步定位，通过MAS手段回交转育了光合主效QTL近等基因系(158个BC3F2株系)，再利用目标QTL对应的置换系与背景亲本杂交构建次级分离群体（已获得BC1F1目标单株37个）；应用高通量转录组测序技术获得SNP位点和对光响应显著基因，利用全基因组测序技术（Solexa技术）进行目标区段的序列分析以获得序列差异的预测基因和新标记位点，选取区段内SNP位点设计开发新标记，再对F2群体PR进行2季测定，进而对其精细定位；利用生物信息学分析、比较基因组分析技术结合PR主效QTL定位结果确定候选基因；再对候选基因进行表达谱分析和初步的功能分析，发掘出水稻高光效基因，为水稻高光效基因遗传机理研究和分子标记辅助育种奠定基础。

本研究在初步定位的基础上，沿用珍汕97B（indica）×IRAT109（japonica）杂交构建的重组自交系后代群体，通过与母本多次回交，辅以分子标记选择，构建染色体片段代换系群体，以携带第10 号染色体目标区段来源于IRAT109，珍汕97B为背景的的材料，进行目标区段内的光合QTL的精细定位并结合其转录组的差异表达显著基因的功能分析来确定光合速率候选基因。.本研究结论如下：.1.目标片段内的分子标记加密，在原目标区段RM596-RM271内加入了8个新的InDel分子标记，标记之间的物理距离缩小到200kb。.2.在构建的CSSL群体中，通过分子标记辅助选择，筛选目标片段内不同位点发生交换重组的纯合单株，进行光合测定和抽穗期调查，发现在目标区段的附近有一已报道的抽穗期基因EHD1，影响群体的抽穗期，为群体的光合QTL精细定位准确性增加难度。该基因来源于IRAT109，表现为抽穗期延迟，母本纯合片段单株的抽穗期均有所提前，而父本纯合单株的抽穗期相对较迟，这一性状与位点的连锁表现对表型检测产生干扰。根据表型结合基因型数据分析，我们初步确定了在GH16-GH18内约为200kb的片段与光合作用相关。 .3.对BC4F1的5个样本2组光暗处理，L1、L2、L3、DK1和DK3样品转录组分析，数据表明，在目标区间内，找到4个差异表达显著的基因：LOC_Os10g30162.1编码淀粉合酶Ⅱ或叶绿体前体，LOC_Os10g30156.1编码热激蛋白Hsp20蛋白结合域，LOC_Os10g31320.1编码尿苷二磷酸葡萄糖醛酸/尿苷二磷酸葡萄糖转移酶蛋白家族，LOC_Os10g30560.1功能未知。在携带有供体IRAT109片段的材料L1、L2和DK1中编码淀粉合酶Ⅱ或叶绿体前体和编码热激蛋白Hsp20蛋白结合域基因的表达都是显著低于珍汕97B，而编码尿苷二磷酸葡萄糖醛酸/尿苷二磷酸葡萄糖转移酶蛋白家族基因表达却有显著提高。.4.转录组差异表达基因功能分析表明，导入目标片段的转录本在捕光色素复合体（Light harvesting chlorophyll protein complex）和植物的昼夜节律（Circadian rhythm)）等途径的功能表达差异显著。