水稻糯性突变体r162的鉴定与基因定位  

Identification and gene mapping of rice glutinous mutant r162

作　　者：何羽喆 徐善斌 陈彦宇 杨雪 段二超 滕烜[1] 王益华[1] 董慧 万建民 HE Yuzhe;XU Shanbin;CHEN Yanyu;YANG Xue;DUAN Erchao;TENG Xuan;WANG Yihua;DONG Hui;WAN Jianmin(State Key Laboratory of Crop Genetics and Germplasm Enhancement/Jiangsu Collaborative Innovation Center for Modern Crop Production,Nanjing Agricultural University,Nanjing 210095,Jiangsu,China;National Key Facility for Crop Gene Resources and Genetic Improvement,Institute of Crop Sciences,Chinese Academy of Agricultural Sciences,Beijing 100081,China)

机构地区：[1]南京农业大学作物遗传与种质创新国家重点实验室/江苏省现代作物生产协同创新中心,江苏南京210095 [2]中国农业科学院作物科学研究所,农作物基因资源与基因改良国家重大科学工程,北京100081

出　　处：《浙江大学学报（农业与生命科学版）》2024年第3期368-381,共14页Journal of Zhejiang University:Agriculture and Life Sciences

基　　金：江苏省重点研发计划项目(BE2022383)。

摘　　要：直链淀粉是影响稻米食味品质和外观品质的重要因素,主要由Waxy(Wx)基因编码的颗粒结合淀粉合酶Ⅰ(granule-bound starch synthaseⅠ, GBSSⅠ)介导合成。本研究从宁粳2号的甲基磺酸乙酯(ethyl methanesulfonate,EMS)诱变材料中筛选到了一个糯性突变体r162,与野生型相比,其籽粒外观呈不透明的云雾状,种子和糙米的粒长、粒宽以及千粒重显著下降。通过扫描电镜对淀粉形态进行观察,发现突变体的淀粉颗粒上存在孔洞。半薄切片碘染结果表明,突变体r162发育时期胚乳的造粉体排列结构与野生型相似,但碘染后颜色更浅。理化性质分析发现,突变体r162的总淀粉、总蛋白质和总脂肪含量与野生型相比无显著差异,但是表观直链淀粉含量极显著下降、胶稠度极显著增加,糊化特性发生明显改变,并且淀粉的崩解值、回生值、回复值以及米粉在尿素中的膨胀体积均小于野生型。通过基因定位和测序发现,突变基因Wx位于第6号染色体短臂上,Wx基因第7个内含子的第1个碱基(Int7-1)由鸟嘌呤(G)替换为腺嘌呤(A),造成Wx基因的编码区存在10个碱基的缺失,导致其编码的GBSSⅠ蛋白自第256位氨基酸起出现错误翻译,使蛋白质翻译提前终止,因此将该等位变异命名为Wx-r162。进一步通过蛋白质印迹法检测发现,突变体中的GBSSⅠ蛋白水平极显著下降。酶活性测定也证实,突变体r162中GBSSⅠ活性明显降低。综上所述,Wx-r162基因被视为一个新的等位变异,这种突变导致GBSSⅠ活性下降,进而造成突变体r162表观直链淀粉含量降低和籽粒表现出不透明的表型。Amylose is widely regarded as an important factor affecting the eating quality and appearance quality of rice.It is mainly synthesized by granule-bound starch synthaseⅠ(GBSSⅠ)encoded by the Waxy(Wx)gene.In this study,the glutinous mutant r162 was obtained from an ethyl methanesulfonate(EMS)-treated population of japonica rice cv.Ningjing 2.Compared with wild type(WT),the grains of this mutant showed a significant decrease in length,width and 1000-grain mass,presenting a dull,opaque appearance reminiscent of clouds or fog.Scanning electron microscopy analysis of starch morphology revealed that there were some pores in the starch granules of mutant r162.The iodine staining of semi-thin sections indicated that the amyloplast arrangement in the developing endosperm of mutant r162 was similar to that in the WT,but the staining color was lighter.Physicochemical property analysis revealed that the total starch,total protein and total lipid contents of mutant r162 and WT were not significantly different.However,the apparent amylose content significantly decreased,and the gel consistency increased in mutant r162.Besides,the pasting property was also significantly altered,and the breakdown,setback,consistency,and the swelling volume of rice flour in urea of mutant r162 were lower than WT.Through the gene mapping and sequencing,the mutant gene Wx was localized on the short arm of chromosome 6.The first base of the seventh intron 7(Int7-1)of the Wx gene in the mutant r162 was replaced from guanine(G)to adenine(A),resulting in a deletion of 10 bases in the Wx coding sequence and causing a mistranslation from the 256th amino acid,leading to premature termination of protein translation.Therefore,it was named Wx-r162.Further Western blot analysis revealed a marked decrease in GBSSⅠprotein levels in mutant r162,and enzyme activity assay also showed that the activity of GBSSⅠin the mutant r162 was significantly lower than that in WT.In summary,the Wx-r162 gene is a novel allelic variant whose mutation causes a decrease in

关 键 词：水稻 稻米品质 直链淀粉含量 Waxy(Wx)基因 颗粒结合淀粉合酶Ⅰ 

分 类 号：S331[农业科学—作物遗传育种]