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cry2Ah-vp基因玉米的抗虫性鉴定
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引用本文:李梦桃,李圣彦,汪海,张杰,苍晶,郎志宏.转cry2Ah-vp基因玉米的抗虫性鉴定.植物保护学报,2020,47(1):74-83
DOI:10.13802/j.cnki.zwbhxb.2020.2019043
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作者单位E-mail
李梦桃 东北农业大学生命科学学院, 哈尔滨 150030
中国农业科学院生物技术研究所, 北京 100081 
 
李圣彦 中国农业科学院生物技术研究所, 北京 100081  
汪海 中国农业科学院生物技术研究所, 北京 100081  
张杰 中国农业科学院植物保护研究所, 植物病虫害生物学国家重点实验室, 北京 100193  
苍晶 东北农业大学生命科学学院, 哈尔滨 150030 cangjing2003@163.com 
郎志宏 中国农业科学院生物技术研究所, 北京 100081 langzhihong@caas.cn 
中文摘要:为获得新型抗虫转基因玉米,将通过群体筛选获得的具有抗虫性的转cry2Ah-vp基因玉米VP1-5采用PCR、Southern blot、实时荧光定量PCR(qPCR)、酶联免疫吸附测定(ELISA)等方法进行阳性植株鉴定、拷贝数分析、转录水平和翻译水平分析,同时通过室内和田间生物活性测定鉴定转基因玉米VP1-5对东方黏虫Mythimna separata和亚洲玉米螟Ostrinia furnacalis的抗性。结果表明,在转基因玉米VP1-5中cry2Ah-vp基因已整合到玉米基因组,以单拷贝的形式插入;cry2Ah-vp基因在转基因玉米VP1-5不同部位组织中均可以正常转录,在灌浆期叶片中的mRNA表达量最高,相对表达量为32.67,在灌浆期穗轴中的mRNA表达量最低,相对表达量为3.74;Cry2Ah-vp蛋白在转基因玉米VP1-5的6叶期各组织中表达量均较高,其中在叶片中的表达量达到2 155.18 ng/g FW,在抽雄期花丝中的表达量最高,达到2 165.86 ng/g FW;且转基因玉米VP1-5对东方黏虫有很高的杀虫活性,接虫3 d后幼虫死亡率达到100.00%;对亚洲玉米螟幼虫也有明显的生长抑制作用。表明转基因玉米VP1-5可作为玉米抗虫育种和害虫防治的种质资源。
中文关键词:害虫防治  苏云金芽胞杆菌  cry2Ah-vp基因  转基因玉米  东方黏虫  亚洲玉米螟
 
Identification of insect resistance in the transgenic maize harboring cry2Ah-vp gene
Author NameAffiliationE-mail
LI Mengtao College of Life Sciences, Northeast Agricultural University, Harbin 150030, Heilongjiang Province, China
Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China 
 
LI Shengyan Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China  
WANG Hai Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China  
ZHANG Jie State Key Laboratory for Biology of Plant Diseases and Insects Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China  
CANG Jing College of Life Sciences, Northeast Agricultural University, Harbin 150030, Heilongjiang Province, China cangjing2003@163.com 
LANG Zhihong Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China langzhihong@caas.cn 
Abstract:To obtain the novel insect-resistant maize, the insect-resistant transgenic event VP1-5 harboring cry2Ah-vp gene was detected for the foreign gene insertion, copy number, transcriptional and translational expression of cry2Ah-vp gene by using PCR, Southern blot, real-time fluorescence quantitative PCR (qPCR) and enzyme-linked immunosorbent assay (ELISA), and the bioassay in the laboratory and field was conducted to identify the resistance of VP1-5 transgenic maize to Mythimna separata and Ostrinia furnacalis. The results showed that cry2Ah-vp gene was integrated into maize genome with a single copy in transgenic maize VP1-5. The cry2Ah-vp gene was correctly transcribed in different tissues showing the highest expression in leaf and the lowest expression in cob during grain-filling stage, and the relative mRNA level were 32.67 and 3.74, respectively. The Cry2Ah-vp protein was expressed relatively high in all tissues at six-leaf stage, especially in leaf where the expression was 2 155.18 ng/g FW. Overall, the highest expression of Cry2Ah-vp protein was 2 165.86 ng/g FW in silk at tassel stage. VP1-5 transgenic maize was demonstrated to have high insecticidal toxicity towards armyworm by the fact that the mortality of armyworm larvae reached 100.00% 3 d after inoculation; VP1-5 obviously inhibited the weight gain of the corn borer larvae. This study indicated that VP1-5 transgenic maize could be a good germplasm resource for insect resistance and pest control.
keywords:pest control  Bacillus thuringiensis  cry2Ah-vp gene  transgenic maize  Mythimna separata  Ostrinia furnacalis
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