转双Bt基因欧洲黑杨毒蛋白季节变化及抗虫性分析  

SEASINAL VARIATION AND INSECT RESISTANCE ANALYSIS OF TOXIN PROTEIN OF Populus nigra L.WITH DOUBLE Bt GENE

作　　者：张骐 王进茂[1] ZHANG Qi;WANG Jinmao(Agricultural University of Hebei,College of Forestry,Baoding,071000,China)

机构地区：[1]河北农业大学林学院,保定071000

出　　处：《内蒙古农业大学学报（自然科学版）》2021年第4期27-33,共7页Journal of Inner Mongolia Agricultural University(Natural Science Edition)

基　　金：国家自然科学基金项目(31660211)。

摘　　要：利用美国白蛾和柳蓝叶甲对转Cry1Ac-Cry3A-BADH-MtlD基因欧洲黑杨进行抗虫性筛选,对筛选出具有双高抗虫性的5个转基因株系进行外源基因PCR检测,利用酶联免疫吸附法(ELISA)对5个转基因株系叶片Bt毒蛋白含量进行检测,分析5个转基因株系叶片中2个Bt基因在生长季节不同时期的表达情况,对其差异进行分析;检测转基因株系对不同虫龄美国白蛾和柳蓝叶甲的抗性。结果表明:(1)初步筛选出5个高抗虫性的转基因株系1号、7号、9号、11号和12号,其对美国白蛾和柳蓝叶甲1龄幼虫致死率均超过90%以上;(2)PCR检测结果表明5个转基因株系叶片中均检测Cry1Ac-Cry3A基因条带,而对照则未检测到,证明外源基因均已插入到受体杨树基因组中;(3)转基因株系均检测到Bt毒蛋白,而对照则未检测出。从6月至10月,转基因株系叶片中Cry1Ac毒蛋白的表达量总体呈上升趋势,但在6月、7月略有下降;Cry3A毒蛋白的表达量从6月至10月总体呈下降趋势;(4)美国白蛾和柳蓝叶甲随虫龄的增大致死率呈下降趋势,不同转基因株系间抗虫性存在显著差异。The Populus nigra L.’s gene was screened for insect resistance by Hyphantria cunea Drury and Plagiodera versicolora.Five transgenic lines with double high insect resistance were screened for exogenous gene detection.Fluorescence quantitative PCR was used to detect the five transgenic lines;Enzyme-linked immunosorbent assay(ELISA)was used to detect the content of Bt toxin in leaves of 5 transgenic lines;and the expressions of 2 Bt genes in leaves of 5 transgenic lines were analyzed in different periods during the growing season,and the differences were analyzed.Resistance of transgenic lines to Hyphantria cunea Drury and Plagiodera versicolora of different insect ages was tested.The results showed that:(1)five transgenic lines 1,7,9,11 and 12 with high insect resistance were preliminarily selected,and the mortality for the first instar larvae of Hyphantria cunea Drury and Plagiodera versicolora was more than 90%.(2)PCR detection results showed that the Cry1Ac-Cry3a gene bands were detected in the leaves of all the 5 transgenic lines,while the non-transgenic control group was not detected,indicating that all the exogenous genes had been inserted into the genome of the recipient poplar.(3)Bt viral protein was detected in all transgenic lines,but not in the control group.From June to October,the expression of toxoprotein Cry1Ac in the leaves of transgenic lines showed an overall trend of increase,but decreased slightly in june and July.The expression of toxoprotein Cry3A decreased from June to October.(4)The mortality of Hyphantria cunea Drury and Plagiodera versicolora decreased with the increase of insect age.There were significant differences in insect resistance among different transgenic lines.

关 键 词：欧洲黑杨 抗虫性筛选 CRY1AC基因 Cry3A基因 

分 类 号：S722.36[农业科学—林木遗传育种]