小麦光温敏雄性不育系337S不育相关基因的遗传转化研究

发布时间：2018-04-21 11:26

本文选题：普通小麦 + 雄性不育　；参考：《华中农业大学》2017年硕士论文

【摘要】：小麦337S不育系是对长日高温、短日低温均敏感的两极光温敏小麦雄性不育系,具有不育性稳定、制种区域广之特点。本实验室早期以337S光温敏雄性不育系在短日低温不育环境条件及正常可育环境条件下的花药为材料,采用数字基因表达谱技术对其进行RNA-Seq测序分析,获得了大量与小麦生殖发育相关的差异基因,并对其中差异表达显著的关键基因进行了分子克隆。同时利用Gateway技术构建了候选基因TaMS337S-1和TaMS337S-2的超量表达载体,通过基因枪和农杆菌介导的转基因技术转化不同小麦品种进行功能验证,探究TaMS337S-1和TaMS337S-2基因在小麦生长过程中的主要作用,并为337S光温敏不育系短日低温败育分子机理的研究提供线索。现已基本完成了这两个基因的遗传转化工作,主要实验结果如下:以华麦2152和华麦2566为受体通过基因枪轰击法转化TaMS337S-1基因,没有得到转基因植株。以华麦2152、华麦2566、郑麦9023、陇春23、科农199和Bob White幼胚为受体材料,通过基因枪轰击转化TaMS337S-2基因得到再生小麦51株,分别为4株华麦2566,41株陇春23,1株Bob White。PCR初步检测后显示2株陇春23为阳性植株,其余均为阴性再生苗。以华麦2152、华麦2566、陇春23和科农199为受体材料,利用农杆菌侵染法转化TaMS337S-1基因得到再生植株7株,分别为3株华麦2566,4株陇春23。PCR结果显示有2株华麦2566为转基因阳性植株,其表型为花丝外露、无花药。以华麦2152、华麦2566、陇春23和科农199为受体材料通过农杆菌转化TaMS337S-2基因共得到9株再生苗,分别为1株华麦2566,4株科农199,4株陇春23,但PCR检测没有发现转基因阳性苗。
[Abstract]:Wheat 337s male sterile line is sensitive to long day high temperature and short day low temperature sensitive male sterile line of wheat, which has stable sterility and wide seed production area. The anthers of 337S photo-thermo-sensitive male sterile lines were used as materials in short day low temperature sterility environment and normal fertile environment in our laboratory. Digital gene expression technique was used to analyze the RNA-Seq sequence of the male sterile lines. A large number of differentially expressed genes related to wheat reproductive development were obtained and the key genes with significant differentially expressed genes were cloned. At the same time, the overexpression vectors of candidate genes TaMS337S-1 and TaMS337S-2 were constructed by using Gateway technology, and then transformed into different wheat varieties by gene gun and Agrobacterium tumefaciens. To explore the main role of TaMS337S-1 and TaMS337S-2 genes in wheat growth, and to provide clues for the study of the molecular mechanism of short-day and low-temperature abortion of 337S photo-thermosensitive male sterile lines. The main results are as follows: Huamai 2152 and Huamai 2566 were used as receptors to transform TaMS337S-1 gene by bombardment of gene gun, and no transgenic plants were obtained. The immature embryos of Huamai 2152, Huamai 2566, Zhengmai 9023, Longchun 23, Konong 199 and Bob White were used as recipient materials, and 51 regenerated wheat plants were obtained by bombardment of TaMS337S-2 gene with gene gun. The results of Bob White.PCR analysis showed that 2 Longchun 23 plants were positive plants, and the rest were negative regenerated plantlets. Using Huamai 2152, Huamai 2566, Longchun 23 and Konong 199 as receptor materials, 7 regenerated plants were obtained by Agrobacterium tumefaciens infection method. The results showed that 2 Huamai 2566 plants were transgenic positive plants. Its phenotype is filaments exposed, no anther. Using Huamai 2152, Huamai 2566, Longchun 23 and Konong 199 as receptor materials, 9 regenerated seedlings were obtained by Agrobacterium tumefaciens transformation into TaMS337S-2 gene, one of them was 1 Huamai 2566 / 4 Konong 1994 strain Longchun 23, but no transgenic seedlings were found by PCR test.
【学位授予单位】：华中农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S512.1

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