水稻巨大胚突变体N2-52垩白形成机制研究  

Study on the mechanism of chalkiness formation in the rice giant embryo mutant N2-52

作　　者：葛鑫源 刘世家[1] 李欣[1] 燕海刚 王永祥 傅玉双 单壮壮 张文伟[1] 王益华[1] 江玲[1] GE Xinyuan;LIU Shijia;LI Xin;YAN Haigang;WANG Yongxiang;FU Yushuang;SHAN Zhuangzhuang;ZHANG Wenwei;WANG Yihua;JIANG Ling(Key Laboratory of Biology,Genetics and Breeding of Japonica Rice in Mid-lower Yangtze River,Ministry of Agricultureand Rural Affairs/The Yangtze River Valley Hybrid Rice Collaboration Innovation Center/Jiangsu CollaborativeInnovation Center for Modern Crop Production,Nanjing Agricultural University,Nanjing 210095,China)

机构地区：[1]南京农业大学农业农村部长江中下游粳稻生物学与遗传育种重点实验室/长江流域杂交水稻协同创新中心/江苏省现代作物生产中心,江苏南京210095

出　　处：《南京农业大学学报》2021年第2期217-224,共8页Journal of Nanjing Agricultural University

基　　金：国家转基因生物新品种培育重大专项(2016ZX08001006);江苏省农业农村厅重大品种选育项目(PZCZ201701)。

摘　　要：[目的]本文旨在通过对水稻巨大胚突变体进行表型分析和基因图位克隆,阐明垩白形成的分子机制。[方法]以粳稻品种‘宁粳2号’及其巨大胚乳突变体N2-52为材料,分析野生型和突变体的农艺性状和籽粒的理化特性,观察N2-52垩白区域的造粉体发育和淀粉颗粒形态;构建N2-52与籼稻品种‘N22’杂交的F_(2)群体,进行突变基因的图位克隆;利用蛋白免疫印迹测定淀粉合成相关酶在发育中种子、成熟种子及其垩白部分的蛋白积累量;测定蔗糖合酶的酶活性。[结果]突变体N2-52籽粒表现为巨大胚、高垩白,其结实率、千粒重极显著低于野生型,粒厚、脂肪含量显著低于野生型,株高、垩白粒率、直链淀粉含量极显著高于野生型。扫描电镜观察表明,N2-52垩白部分淀粉颗粒呈疏松的不规则排列,半薄切片也证实该部分造粉体分布疏松,且胚乳细胞内存在空泡。RVA(rapid visco analyzer)和尿素膨胀试验表明,N2-52的淀粉理化性质发生改变。突变基因被定位于第7染色体短臂的1个42 kb的区域内,测序结果表明该区域中只有1个已知基因Os07g0603700发生突变。Western blot结果显示,淀粉合成相关酶在垩白部分的蛋白积累量发生变化。N2-52的蔗糖合酶活性显著高于野生型。[结论]Os07g0603700基因的突变影响淀粉合成相关基因的表达,对淀粉的合成产生重要影响,推测其导致垩白的产生。[Objectives]This study aimed to elucidate the molecular mechanism by which the giant embryo gene caused chalky endosperm,based on the phenotypic analysis and gene positional-cloning of a rice giant embryo mutant.[Methods]The agronomic traits of the japonica rice variety‘Ningjing 2’(the wild-type)and the giant embryo mutant N2-52 were analyzed.The physicochemical characteristics of mature grains,and the amyloplast development and starch granule morphology in chalky areas of N2-52,were also examined.The F_(2) population of N2-52 and the indica rice variety‘N22’was used for mapping the mutant gene.The amounts of starch synthetic enzymes were examined in developing and mature seeds,and the chalky portions,by Western blot.The enzyme activity of sucrose synthase was determined.[Results]The mutant grains showed giant embryo and high chalkiness rate.Compared with the wild-type,N2-52 had very significantly lower seed setting rate and 1000-grain weight,significantly lower grain thickness and fat content,and significantly higher plant height,chalky rice rate and amylose content.Scanning electron microscopy showed that the starch granules in the chalky region of N2-52 were loosely and irregularly arranged.Semi-thin sections also indicated that the amyloplasts in this region were scattered in the cytosol with large air spaces.RVA(rapid visco analyzer)analysis and urea swelling assays showed that the physicochemical properties of N2-52 starch were altered.The mutated gene was finally located in a 42 kb region on the short arm of chromosome 7,and sequencing analysis revealed that a known gene,Os07g0603700,was mutated in N2-52.Western blot analysis showed that the amounts of starch synthetic enzymes in the chalky parts of N2-52 grains were changed.The activity of sucrose synthase of N2-52 was significantly higher than that of the wild type.[Conclusions]The mutation of Os07g0603700 gene affected the expression of starch synthetic genes and finally the synthesis of starch,which possibly led to chalkiness.

关 键 词：水稻 垩白 巨大胚 造粉体 突变体 精细定位 

分 类 号：S511.032[农业科学—作物学]