西瓜花叶病毒（WMV）外壳蛋白基因的转化及其抗病分析

发布时间：2018-06-09 23:20

  本文选题：伽师瓜 + 植物病毒病　； 参考：《新疆大学》2017年硕士论文

【摘要】：甜瓜(Cucumis melo L.)不仅高产、优质,而且经济效益高,其市场销售范围也很广,在新疆栽培历史悠久,也是世界重要的经济作物。素有“植物癌症”之称的植物病毒病,是影响多种农作物减产的主要原因,在瓜类作物中造成的危害尤为严重。病毒病一旦大面积发生,甜瓜就会严重减产,品质下降,甚至绝收。而且病毒在田间还会经常发生复合侵染,这使得病毒病的防治更加困难。近几年新疆瓜果产业一直受到植物病毒的影响,而主栽甜瓜品种不抗植物病毒病,所以利用基因工程技术转化病毒来源的外壳蛋白基因也是一种可行的方法,由此开展实验并得到以下结果:1.从新疆吐鲁番收集的发病叶片,分离纯化后确定为西瓜花叶病毒。克隆获得西瓜花叶病毒外壳蛋白(coat protein,CP)基因序列,经HindⅢ与XbaⅠ酶切后插入pCAMBIA2300载体中,获得植物双元表达载体。2.以厚皮甜瓜伽师为材料,建立以子叶为外植体的快速高效再生体系。甜瓜子叶外植体接种在MSB+1.5 mg·L-1 6-BA诱导分化培养基上,诱导率可达75%;添加不同浓度AgNO3诱导不定芽,以MSB+1.5 mg·L-1 6-BA+2.0 mg·L-1AgNO3最适,诱导率可达65%,产生单芽较多,缩短了再生周期。3.通过农杆菌GV3101的介导甜瓜和本氏烟草,得到了许多拟阳性转基因烟草和甜瓜,提取拟阳性植株基因组DNA,经过PCR鉴定,得到了11株转基因烟草,4株转基因甜瓜。4.对转基因烟草的SOD、POD和CAT的酶活性进行检测,发现这三种酶的活性均高于野生型烟草,初步验证了转基因烟草对西瓜花叶病毒存在抗性,可作为早期筛选抗性植株的辅助指标。统计T1代幼苗Kan抗性遗传分离比为3:1的转基因烟草,得到1、4、5、6、9、10、11这七个符合株系。本实验结果为进一步开展病毒病与植物抗病性的研究奠定基础。
[Abstract]:Cucumis melo L. Not only high yield, good quality, but also high economic benefit, its marketing range is also very wide, it has a long history of cultivation in Xinjiang, and is also an important cash crop in the world. Plant virus disease, known as "plant cancer", is the main reason that affects the yield reduction of many crops, especially in melon crops. Once the virus disease occurs in a large area, melon will seriously reduce production, quality decline, or even end-harvest. And the virus often occurs in the field complex infection, which makes it more difficult to prevent and cure the virus disease. In recent years, the melon and fruit industry in Xinjiang has been affected by plant viruses, and the main melon varieties are not resistant to plant virus disease, so it is also a feasible method to use genetic engineering technology to transform the coat protein gene from the virus source. The experiment was carried out and the following results were obtained: 1: 1. Watermelon mosaic virus was isolated and purified from the infected leaves collected from Turpan, Xinjiang. The coat protein (CPP) gene of watermelon mosaic virus was cloned and inserted into pCAMBIA2300 vector by Hind 鈪，

本文编号：2001177