一份自然突变雄性核不育水稻材料的遗传分析与基因定位
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摘要
作物育性是作物的重要农艺性状之一,是作物杂种优势利用的关键因素。而水稻雄性不育是自然界的普遍现象,水稻雄性不育的研究对水稻的育种具有重要利用价值。本研究利用从D295B与昌丰B杂交F_4代中自然突变得到的一雄性不育突变植株,定名为SC-ms-2,将其与花B等水稻材料杂交,对SC-ms-2进行育性研究、细胞形态学研究和遗传分析,并利用分子标记结合分离群体分组分析法对控制此性状的雄性不育基因进行了基因精细定位,主要结果如下:
1、SC-ms-2花药白色、透明、细小,成熟的花药具有不育花粉粒,属于有花粉型雄性不育系,不育性表现稳定,不受光温条件影响;其颖花的雌蕊等其它花结构发育正常。与可育品种材料杂交,F_1代正常可育,F_2代可育与不育植株数表现为3:1分离比例,表明此雄性不育性状受一对隐性核基因控制,不受细胞质遗传的影响。
2、石蜡切片细胞学观察,在小孢子时期之前,SC-ms-2突变体和野生型花粉都发育正常,无明显区别。在进入小孢子发育时期后,野生型绒毡层细胞可正常浓缩、降解,而突变体则出现小孢子发育呈畸形,绒毡层加厚并延迟降解,不能发育成正常成熟花粉粒。
3、遗传效应分析表明:SC-ms-2生育期84天,株高84cM,有效穗10.3个,穗长24.6cM,平均着粒数176颗,包颈率7.34%,包颈度为26.65%,比对应的近等基因系杂合可育株系生育期短0.67天,株高低13.39cM,有效穗多1.1个,穗长长3.8cM,平均着粒数多38.4颗。
4、根据遗传分析的结果,将SC-ms-2/花B的F_2代作为定位群体,采用分离群体分组分析法,利用SSR标记进行初步定位,初步定位在水稻9号染色体上RM34431与RM3600之间,为进一步进行精细定位,更换花B群体为亲本差异更大的日本晴群体,分析在RM24431与RM3600之间SSR分子标记在亲本之间多态性,最终把SC-ms-2不育基因精细定位到分子标记RM24451、RM7048之间,遗传图距分别为0.3 cM和0.6 cM;在这两个标记间约172KB的区域内找到25个已经预测的候选基因,经分析,编码雄性不育蛋白的基因LOC_Os09g27620最有可能为目标基因。经检索,该隐性不育基因属首次利用分子标记定位的新基因,暂命名为ms92(t)。
Crop fertility character was one of the most important agronomic traits of crop,and was key factor of the utility of heterosis in hybrid crop.The male sterility of rice was a common phenomenon occasionally occurring in plant,and the study of male sterility of rice had important utilizing value of rice breeding.In this research,the spontaneous rice mutant with male sterility,SC-ms-2,was obtained from the F4 progenies of the cross D 297B X Changfeng B.The cross was between SC-ms-2 and Hua B along with some other rice materials.The fertility character,morphological cytology and genetic analysis were studied by us.At the same time,with using bulked segregation analysis,the recessive genic male sterility gene was fine mapping by SSR molecular maker.The main results are summarized as bellows:
1.The anther of SC-ms-2 was white,transparent,gracile and produced male sterility pollen.It belonged to pollen type.The sterility character was stable,not affected by temperature and light change,but the pistil of glume and other flower structure were normal.Furthermore,all of the F_1 plants from crosses between this mutant and wild type were male fertile.The fertility and male sterility were segregated in all the F_2 populations from the three crosses and the segregation ratios fit well to 3:1.These results indicated that male sterility phenotype of this mutant SC-ms-2 was controlled by a single recessive genic gene,which didn't affect by cytoplasm.
2.By observation of paraffin section,SC-ms-2 and wild type had no differentiation in the microspore mother cell stage,prophase stage of meiosis and tetrad stage before microspore morphogenesis.After the pollens developed in the microspore morphogenesis, the tapetum of wild type concentrated and disassembled normally,but the microspore of male sterility mutant were oafish,the tapetum cells was incrassated,at last microspore couldn't be develop normal pollens.
3.Comparing with heterozygote of near-isogenetic lines,the result of genetic effect analysis showed that the growth duration of SC-ms-2 had 84 days,0.67 days more than heterozygote;the plant height had 84 cM,13.39cM lower than heterozygote;the valid spikes were 10.3,1.1 more than heterozygote;the panicle length was 24.6cM,3.8cM longer than heterozygote;the spikelets per panicle had 176 grains,38.4 grains more than heterozygote;at the same time,the panicle enclosure rate of SC-ms-2 was 7.34%,and the panicle enclosure length rate was 26.65%.
4.According to genetic analysis,by using bulked segregation analysis on an F_2 population developed from the cross of SC-ms-2 and Hua B,this gene was mapped close to two simple sequence repeat(SSR) makers,RM24431 and RM3600,on chromosome 9. Further,fine mapping was carried out by using the F_2 population developed from the cross between MS-sc-2 and Nipponbare,and this gene was located between SSR molecular maker RM24451 and RM7048,with genetic distance of 0.3 cM and 0.6 cM,respectively, and physical distance approximately 172 kb.Bioinformatics analysis showed that LOC_Os09g27620 maybe the target gene.Our results showed that this gene was distinguished from all other genes with male sterility in rice reported,and it was designated ms92(t),temporally.
1、SC-ms-2花药白色、透明、细小,成熟的花药具有不育花粉粒,属于有花粉型雄性不育系,不育性表现稳定,不受光温条件影响;其颖花的雌蕊等其它花结构发育正常。与可育品种材料杂交,F_1代正常可育,F_2代可育与不育植株数表现为3:1分离比例,表明此雄性不育性状受一对隐性核基因控制,不受细胞质遗传的影响。
2、石蜡切片细胞学观察,在小孢子时期之前,SC-ms-2突变体和野生型花粉都发育正常,无明显区别。在进入小孢子发育时期后,野生型绒毡层细胞可正常浓缩、降解,而突变体则出现小孢子发育呈畸形,绒毡层加厚并延迟降解,不能发育成正常成熟花粉粒。
3、遗传效应分析表明:SC-ms-2生育期84天,株高84cM,有效穗10.3个,穗长24.6cM,平均着粒数176颗,包颈率7.34%,包颈度为26.65%,比对应的近等基因系杂合可育株系生育期短0.67天,株高低13.39cM,有效穗多1.1个,穗长长3.8cM,平均着粒数多38.4颗。
4、根据遗传分析的结果,将SC-ms-2/花B的F_2代作为定位群体,采用分离群体分组分析法,利用SSR标记进行初步定位,初步定位在水稻9号染色体上RM34431与RM3600之间,为进一步进行精细定位,更换花B群体为亲本差异更大的日本晴群体,分析在RM24431与RM3600之间SSR分子标记在亲本之间多态性,最终把SC-ms-2不育基因精细定位到分子标记RM24451、RM7048之间,遗传图距分别为0.3 cM和0.6 cM;在这两个标记间约172KB的区域内找到25个已经预测的候选基因,经分析,编码雄性不育蛋白的基因LOC_Os09g27620最有可能为目标基因。经检索,该隐性不育基因属首次利用分子标记定位的新基因,暂命名为ms92(t)。
Crop fertility character was one of the most important agronomic traits of crop,and was key factor of the utility of heterosis in hybrid crop.The male sterility of rice was a common phenomenon occasionally occurring in plant,and the study of male sterility of rice had important utilizing value of rice breeding.In this research,the spontaneous rice mutant with male sterility,SC-ms-2,was obtained from the F4 progenies of the cross D 297B X Changfeng B.The cross was between SC-ms-2 and Hua B along with some other rice materials.The fertility character,morphological cytology and genetic analysis were studied by us.At the same time,with using bulked segregation analysis,the recessive genic male sterility gene was fine mapping by SSR molecular maker.The main results are summarized as bellows:
1.The anther of SC-ms-2 was white,transparent,gracile and produced male sterility pollen.It belonged to pollen type.The sterility character was stable,not affected by temperature and light change,but the pistil of glume and other flower structure were normal.Furthermore,all of the F_1 plants from crosses between this mutant and wild type were male fertile.The fertility and male sterility were segregated in all the F_2 populations from the three crosses and the segregation ratios fit well to 3:1.These results indicated that male sterility phenotype of this mutant SC-ms-2 was controlled by a single recessive genic gene,which didn't affect by cytoplasm.
2.By observation of paraffin section,SC-ms-2 and wild type had no differentiation in the microspore mother cell stage,prophase stage of meiosis and tetrad stage before microspore morphogenesis.After the pollens developed in the microspore morphogenesis, the tapetum of wild type concentrated and disassembled normally,but the microspore of male sterility mutant were oafish,the tapetum cells was incrassated,at last microspore couldn't be develop normal pollens.
3.Comparing with heterozygote of near-isogenetic lines,the result of genetic effect analysis showed that the growth duration of SC-ms-2 had 84 days,0.67 days more than heterozygote;the plant height had 84 cM,13.39cM lower than heterozygote;the valid spikes were 10.3,1.1 more than heterozygote;the panicle length was 24.6cM,3.8cM longer than heterozygote;the spikelets per panicle had 176 grains,38.4 grains more than heterozygote;at the same time,the panicle enclosure rate of SC-ms-2 was 7.34%,and the panicle enclosure length rate was 26.65%.
4.According to genetic analysis,by using bulked segregation analysis on an F_2 population developed from the cross of SC-ms-2 and Hua B,this gene was mapped close to two simple sequence repeat(SSR) makers,RM24431 and RM3600,on chromosome 9. Further,fine mapping was carried out by using the F_2 population developed from the cross between MS-sc-2 and Nipponbare,and this gene was located between SSR molecular maker RM24451 and RM7048,with genetic distance of 0.3 cM and 0.6 cM,respectively, and physical distance approximately 172 kb.Bioinformatics analysis showed that LOC_Os09g27620 maybe the target gene.Our results showed that this gene was distinguished from all other genes with male sterility in rice reported,and it was designated ms92(t),temporally.
引文
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