【作者】 张伟；

【导师】 张方东；

【作者基本信息】 华中农业大学 ， 作物栽培学与耕作学， 2023， 硕士

【摘要】 玉米(Zea mays L.)是具有广泛用途的农作物,对我国的民生和经济发展有着至关重要的作用。通过突变体解析玉米基因的功能,对玉米的品种改良有着重要的意义。本实验室前期结合玉米Ac/Ds转座子和超表达标签生成系技术初步建成了玉米超表达突变体生成系统。本研究在此基础上对超表达突变体生成系统进行补充并进行了突变体筛选鉴定。主要研究结果如下:1.创建了三个玉米超表达突变体生成系DATTL-68、DATTL-69、DATTL-70(Ds Based Activation Tagging Transposon Line)。通过雌穗籽粒颜色的变化计算人工转座子的转座频率分别为42.50%、33.10%、37.10%。2.构建了两个超表达突变体转座子生成系:Zm Ds-697-Transposon系和Zm DsRsc-Bar-Transposon系。利用TAIL-PCR和重测序技术对转座子系进行了T-DNA插入位点的鉴定以及侧翼序列的分离,得到了18个转座子系的T-DNA的插入位点。将实验室建立的54个生成系数量增加至72个,并补充了第八号染色体短臂上的转座子生成系,初步建成了玉米超表达突变体生成系统核心群体。3.在2021年至2022年四个播种季节对玉米超表达突变体生成系进行常规农艺条件下的筛选,得到了数百个表型出现变异的突变体植株,涵盖了玉米的株形、叶色、叶片形态、生长势以及雌雄穗的发育形态,其中7个突变性状可以稳定遗传。4.对性状稳定遗传的突变体进行鉴定和候选基因确定,利用重测序技术分离定位宽叶突变体LW(Leaf Width)人工转座子标签插入位点位于玉米基因组第七号染色体chr7:128326215 bp处,插入GRMZM2G095254基因的CDS序列5’端59 bp处。预测了四个候选基因:GRMZM2G095254、GRMZM2G308503、GRMZM2G396350、GRMZM2G161619。本研究对玉米超表达突变体生成系统进行了补充完善,并进行了超表达突变体的筛选,选择其中能够稳定遗传的突变体进行研究,预测了候选基因,为玉米超表达突变体生成系统的应用探索出了规范化的流程,推进了玉米大型超表达突变体库的构建进程。更多还原

【Abstract】 Maize(Zea mays L.)is a widely used crop,and plays a very important role in the people’s livelihood and economic development.The function analysis of maize genes by mutants is of great significance for maize variety improvement.In our laboratory,we preliminatively built a maize overexpression mutant generation system by combining maize Ac/Ds transposons and overexpression tag generation line technology.In this study,the overexpression mutant generation system was supplemented and mutant screening and identification were carried out.The main research results are as follows:1.Three maize overexpression mutant lines DATTL-68,DATTL-69 and DATTL-70 were created.The transposition frequencies of artificial transposons were 42.50%,33.10% and 37.10%,respectively.2.Two transposon lines were constructed: Zm Ds-697-Transposon line and Zm DsRsc-Bar-Transposon line.TAIL-PCR and resequencing techniques were used to identify TDNA insertion sites of transposon lines and isolate flank sequences,and T-DNA insertion sites of 18 transposon lines were obtained.The number of the 54 lines established in the laboratory was increased to 72,and the transposon lines on the short arm of chromosome 8were added,and the core population of maize overexpression mutant generation system was initially established3.During the four sowing seasons from 2021 to 2022,the overexpressed mutant lines of maize were screened under conventional agronomic conditions,and hundreds of mutant plants with phenotypic variation were obtained,covering plant shape,leaf color,leaf morphology,growth potential and the developmental morphology of male and female ears of maize.Among them,7 mutant characters could be inherited stably.4.The mutant with stable character inheritance was identified and the candidate gene was determined.Resequencing technology was used to isolate and locate the LW(Leaf Width)artificial transposon tag insertion site located on chromosome 7 of maize genome:At 128326215 bp,insert the 5’end of the CDS sequence of GRMZM2G095254 gene at 59 bp.Four candidate genes were predicted: GRMZM2G095254,GRMZM2G308503,GRMZM2G396350,GRMZM2G161619.In this study,the generation system of maize overexpression mutants was supplemented and improved,and the overexpression mutants were screened.The mutants with stable inheritance were selected for study,candidate genes were predicted,and a standardized process was explored for the application of the generation system of maize overexpression mutants,which promoted the construction process of large maize overexpression mutants library.更多还原