水稻粒宽突变体gw87的基因定位及候选基因分析  

Gene Mapping and Candidate Gene Analysis of Grain Width Mutant gw87 in Rice

作　　者：张翔宇 郭佳 王三 陈聪萍 孙昌辉[1] 邓晓建[1] 王平荣[1] ZHANG XiangYu;GUO Jia;WANG San;CHEN CongPing;SUN ChangHui;DENG XiaoJian;WANG PingRong(Rice Research Institute,Sichuan Agricultural University,Chengdu 611130)

机构地区：[1]四川农业大学水稻研究所,成都611130

出　　处：《中国农业科学》2021年第12期2487-2498,共12页Scientia Agricultura Sinica

基　　金：国家自然科学基金(31971869)、国家重点研发计划(2017YFD0100201)。

摘　　要：【目的】对水稻粒宽突变体gw87(grain width 87)进行表型鉴定、遗传分析、基因定位及候选基因分析,为探明该基因调控水稻籽粒大小的分子机制及应用潜力奠定基础。【方法】利用甲基磺酸乙酯诱变籼稻恢复系材料676R,获得一份籽粒宽度和千粒重显著增加的突变体gw87。对该突变体进行表型观察、农艺性状调查及外源油菜素内酯(BL)敏感性、叶绿素含量、光合参数测定,了解其表型特征及生理特性。调查gw87与676R杂交F1的表型和F2群体的分离情况,分析其遗传行为;选取该群体中的突变植株进行高通量测序,并利用gw87与粳稻品种日本晴杂交的F2代作为定位群体,通过MutMap分析和分子标记定位,遴选候选基因并进行DNA和cDNA测序验证。利用qRT-PCR分析gw87和676R中BR合成途径基因OsDWARF4、D11和D2的表达差异。【结果】与野生型亲本676R相比,gw87突变体的株高降低,节间缩短,其中,倒一节间长度缩短最大,并呈现出扭曲的形态;叶片长度减少,宽度增加;单株有效穗数、主穗穗长和结实率显著降低,但籽粒宽度和千粒重显著增大。BL敏感性试验显示,gw87突变体幼苗对外源BL的敏感性降低。光合色素和光合参数测定表明,gw87光合色素含量增加,光合速率也有所增加。遗传分析表明gw87的突变性状是由1对隐性核基因控制。MutMap分析显示gw87突变基因位于第5染色体中部,在该染色体区域仅有1个碱基突变引起编码氨基酸变化;分子标记连锁分析表明该突变基因位于InDel标记X2和X3之间约101 kb的染色体区域;综合这两方面分析结果,最后遴选出gw87候选基因是编码一个含有AP2/EREBP DNA绑定结构域的转录因子基因LOC_Os05g32270。对该候选基因进行DNA和cDNA测序验证,发现gw87突变体中该基因DNA的第1041位的碱基由G突变为A,导致与该位点相邻的76 bp内含子序列被剪接为外显子,引起阅读框移码,蛋白翻译提前终�【Objective】Phenotypic identification,genetic analysis,gene mapping and candidate gene analysis of grain width mutant gw87(grain width87)were performed in rice,so as to lay a good foundation for further understanding molecular mechanism and utilization potentiality of this gene regulating rice grain size.【Method】Through ethyl methanesulfonate mutagenesis,we isolated gw87 mutant showing significant increase of grain width and 1000-grain weight from indica restorer line material 676R.To identify phenotypic and physiological characteristics of the mutant,we performed observation of phenotypes,investigation of agronomic traits,sensitivity analysis of exogenous brassinolide(BL),determination of chlorophyll contents and photosynthetic parameters by using gw87 and its wild type.To analyze its genetic behavior,we investigated F1 phenotypes and F2 segregation of the crossing combination between gw87 and its wild-type parent 676R.To find candidate gene of gw87,we conducted high-throughput sequencing and MutMap analysis of 30 mutant plants from the F2 population of gw87×676R,and molecular mapping using the F2 population crossing gw87 and japonica cultivar Nipponbare.Meanwhile,we confirmed the candidate gene by sequencing its DNA and cDNA sequences.In addition,we detected expression levels of brassinolide synthesis pathway genes OsDWARF4,D11 and D2 using qRT-PCR analysis.【Result】Compared with the wild-type parent 676R,the gw87 mutant showed significant reduction of productive panicle number per plant,main panicle length and seed setting rate,but significant increase of grain width and 1000-grain weight.Meanwhile,the mutant displayed decreased plant height and shortened internodes,in which the first internode was distorted and its length is shortened the most.Besides,leaf length of gw87 was decreased but the width was increased.Sensitivity assays of exogenous BL suggested that the sensitivity of gw87 seedlings to exogenous BL was reduced.Determination of photosynthetic pigments and photosynthetic parameters showed

关 键 词：水稻(Oryza sativa L.) 籽粒 千粒重 油菜素内酯 AP2类转录因子 

分 类 号：S511[农业科学—作物学]