3个抗虫耐盐基因载体对烟草的遗传转化及表达分析  

Genetic Transformation and Expression Analysis of Tobacco by 3 Insectsresistance and Salt-tolerance Genes Transformation Vector

作　　者：肖丹丹[1] 张旭[1] 任亚超[1] 王桂英[1,2] 左力辉 董研[1] 杨敏生[1] 王进茂[1] XiaoDandan Zhang Xu Ren Yachao Wang Guiying Zuo Lihui Dong Yan Yang Minsheng Wang Jinmao(Forestry College, Agricultural University of Hebei, Baoding, 071001 Agriculture and Forestry Sciences of Langfang, Langfang, 065000)

机构地区：[1]河北农业大学林学院,保定071001 [2]廊坊市农林科学院,廊坊065000

出　　处：《分子植物育种》2017年第1期166-173,共8页Molecular Plant Breeding

基　　金：国家自然科学基金项目(31370663);国家"863"计划项目(2013AA102703)共同资助

摘　　要：病虫害和土壤盐渍化是影响烟草生长发育的重要因素,转基因技术已成为改善这一现状的有效手段。本研究将部分改造的Bt基因Cry1Ac基因、Cry3A基因及NTHK1基因构建在同一载体上,利用农杆菌介导法转化野生烟草植株。经PCR检测,获得10个转基因株系。经荧光定量PCR检测,Cry1Ac基因表达量在104~10~5数量级,1号转录丰度1.58×10~5为最高,而Cry3A基因表达量在10~5~10~6数量级,10号转录丰度1.34×10~6为最高;10个转基因株系双Bt基因均得到表达且存在表达差异,而对照烟草没有Bt基因表达。经ELISA检测,转基因烟草Cry1Ac蛋白平均表达量为150.441 ng/g,变化在0.86~201.74 ng/g;Cry3A蛋白平均表达量为1 192.592 ng/g,变化在23.33~2413.47 ng/g。经过对烟草斜纹夜蛾的虫试,初步证明转基因植株获得了抗虫性。组培苗耐盐试验表明,不同盐浓度条件下,转基因株系的分化状况优于对照。证明抗虫耐盐多基因的一次性转化,可提高烟草的抗虫能力和耐盐性。Pests and soil salinization are important factors for affecting the growth of tobacco, transgenic technology has become an effective means of improving this situation. In this study, we used Agrobacterium mediated transformation of Cry1 Ac gene, Cry3 A gene and NTHK1 gene transformed wild tobacco plants, which were detected by PCR, and 10 transgenic lines were obtained.The fluorescence quantitative PCR detection showed, the expression of Cry1 A cgene reached104~10~5magnitude, the highest transcript abundance was up to1.58×10~5. While the expression of Cry3 A gene reached 10~5~10~6magnitude, the highest transcript abundance was up to 1.34 ×10~6, in the 10 transgenic lines, double Bt genes were expressed, and there were differences in the expression, but the control wasn’t detected the expression of double Bt genes. ELISA test showed the average expression of Cry1 Ac was 150.441 ng/g, the expression changed between 0.86 ng/g and 201.74 ng/g; the average expression of Cry3 A was 1192.592 ng/g, the expression changed between23.33 ng/g and 2413.47 ng/g. After the Prodenialitura（Fabricius）test, it showed that transgenic tobacco preliminarily obtained insect resistance. In salt test,transgenic plants growed better than the control condition. It proved that the multi genes were tra nsformed at a time, which improved the insects-resistance and salt-tolerance of tobacco.

关 键 词：烟草 CRY1AC基因 Cry3A基因 NTHK1基因 多基因表达 

分 类 号：S572[农业科学—烟草工业] Q943.2[农业科学—作物学]