小麦不同胚性愈伤率成熟胚特性比较和PvTB1基因转化研究

发布时间：2018-01-01 21:05

本文关键词：小麦不同胚性愈伤率成熟胚特性比较和PvTB1基因转化研究　出处：《西北农林科技大学》2016年硕士论文　论文类型：学位论文

【摘要】：小麦是世界第一大粮食作物,对人类生活有着十分重要的意义。由于全球耕地面积减少,人口数量逐年上升,提高粮食产量刻不容缓。近年来植物转基因技术发展迅速,已在小麦遗传改良中得到了初步应用,为培育高产品种开辟了一条新的途径。目前,在小麦遗传转化的受体材料中,胚性愈伤组织应用最为普遍。成熟胚由于不受生长季节限制、可周年取材且供给充足,被认为是诱导胚性愈伤组织最为方便的外植体。小麦成熟胚的特征特性与胚性愈伤组织诱导的数量、质量必然具有一定相关性,迄今国内外尚未有相关研究的报道。同时,分蘖是小麦重要的农艺性状之一,对小麦高产、稳产影响显著,而采用遗传转化技术对小麦分蘖特性进行遗传改良,将具有重要的育种和生产应用前景。为了探究胚性愈伤率诱导效果不同的普通小麦成熟胚特性的差异,本研究以胚性愈伤率不同的10种小麦基因型的成熟胚为材料,对其部分生理生化指标进行了测定,并观察分析了其外部形态和内部微观结构。同时,利用基因枪法,将克隆自生物能源作物柳枝稷中调控分蘖性状的基因PvTB1对普通小麦进行了遗传转化。取得的主要研究结果如下:1.以胚性愈伤率不同的10种小麦基因型的成熟胚为材料,对胚性愈伤率高、低不同的两类小麦成熟胚的可溶性糖、可溶性蛋白、纤维素、谷胱甘肽和游离氨基酸的测定。结果显示,两类小麦成熟胚的可溶性蛋白和纤维素含量存在显著性差异,而可溶性糖、谷胱甘肽和游离氨基酸的含量在两类小麦成熟胚间的差异不显著。相关性分析发现,小麦成熟胚胚性愈伤率与纤维素含量存在极显著正相关(P=0.002,P0.01),与可溶性蛋白含量存在显著负相关(P=0.01,P0.05)。2.利用体视显微镜对胚性愈伤率不同的10种小麦基因型的成熟胚外部形态进行观察,结果发现胚性愈伤率高、低不同的两类小麦成熟胚的高度之间存在显著性差异。利用石蜡切片方法对成熟胚的内部结构进行观察比较,结果显示两类小麦成熟胚的内部结构之间差异不显著。3.以SQ26、西农928、西农1376和郑麦9023小麦基因型为转化外植体材料,采用基因枪法,将调控柳枝稷分蘖性状的基因PvTB1对普通小麦进行遗传转化,经筛选共获得409株抗性苗,移栽到温室后存活350株。通过PCR鉴定分析,在郑麦9023中获得1株含有目标基因的转基因植株,转化效率约为0.01%。通过对转基因植株的分蘖数、穗长、株高、旗叶长、穗下第一节长和小穗数6个农艺性状进行统计分析,结果表明转基因植株与对照植株之间并没有显著性差异,初步说明Pv TB1基因在普通小麦中没有表达或表达量不足。
[Abstract]:Wheat is the largest food crop in the world, which is of great significance to human life. Due to the decrease of global cultivated land, the population has increased year by year. It is urgent to increase grain yield. In recent years, plant transgenic technology has been developed rapidly and has been applied in wheat genetic improvement, which opens a new way for breeding high-yield varieties. Embryogenic callus is the most common recipient material in wheat genetic transformation. Mature embryos can be obtained annually and supplied with sufficient materials because they are not restricted by growing season. It is considered that it is the most convenient explant to induce embryogenic callus. The characteristic characteristics of wheat mature embryo must have certain correlation with the quantity and quality of embryogenic callus induction. At the same time, tillering is one of the important agronomic traits of wheat, which has a significant effect on high yield and stable yield of wheat. Genetic transformation technology is used to improve the tillering characteristics of wheat. In order to explore the difference of mature embryo characteristics of common wheat with different embryogenic callus induction rate. In this study, some physiological and biochemical indexes of mature embryos of 10 wheat genotypes with different embryogenic callus rates were measured, and their external morphology and internal microstructure were observed and analyzed. Using genetic gunshot. The genetic transformation of common wheat was carried out by cloning the gene PvTB1, which was cloned from switchgrass, a bioenergy crop, to regulate tillering traits. The main results were as follows:. 1. The mature embryos of 10 wheat genotypes with different embryogenic calli were used as materials. The determination of soluble sugar, soluble protein, cellulose, glutathione and free amino acid in two kinds of wheat mature embryos with high and low embryogenic callus rate. There was significant difference in soluble protein and cellulose content between the two types of wheat mature embryos, while the contents of soluble sugar, glutathione and free amino acids were not significantly different between the two types of wheat mature embryos. There was a very significant positive correlation between the callus rate of mature embryo and cellulose content, and a significant negative correlation between callus rate and soluble protein content (P0. 01). The external morphology of mature embryos of 10 wheat genotypes with different embryogenic calli was observed by stereoscopic microscope. The results showed that the embryogenic callus rate was high. There were significant differences in the height of mature embryos between the two types of wheat. The internal structure of mature embryos was observed and compared by paraffin section method. The results showed that there was no significant difference in the internal structure of mature embryos between the two types of wheat. The genotypes of SQ26, Xinong 928, Xinong 1376 and Zhengmai 9023 were used as the transformed explants. The gene PvTB1, which regulates the tiller character of switchgrass, was transformed into common wheat by gene gun method, and a total of 409resistant seedlings were obtained by screening. A transgenic plant containing target gene was obtained from Zhengmai 9023 by PCR analysis. The transformation efficiency was about 0.01. The agronomic characters of transgenic plants including tiller number, ear length, plant height, flag leaf length, first node length under ear and spikelet number were analyzed statistically. The results showed that there was no significant difference between transgenic plants and control plants. The results showed that PV TB1 gene was not expressed or was not expressed in common wheat.
【学位授予单位】：西北农林科技大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：S512.1

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