| 转G2-EPSPS基因玉米D-3侧翼序列分析与转化体特异性检测方法 |
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| 引用本文： | 郭翠,张维,余桂容,周正富,李亮,冯帅,陈明,王劲.转G2-EPSPS基因玉米D-3侧翼序列分析与转化体特异性检测方法[J].作物杂志,2016,32(1):69-20. |
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| 作者姓名： | 郭翠 张维 余桂容 周正富 李亮 冯帅 陈明 王劲 |
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| 作者单位： | 1 西南科技大学生命科学与工程学院,621010,四川绵阳2 中国农业科学院生物技术研究所,100081,北京3 四川省农业科学院生物技术核技术研究所,610066,四川成都 |
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| 基金项目： | 转基因重大专项(2014ZX0800301B);国家自然科学基金(31370126);国家“973”计划(2013CB733903);国家“863”计划(2012AA063503) |
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| 摘 要： | 通过一次3'端高效热不对称交互PCR（hiTAIL-PCR）和一次长链PCR扩增方法获得了转抗草甘膦基因（G2-EPSPS）玉米品系D-3的外源DNA插入片段的全DNA序列（T-DNA）及两端侧翼序列,并建立了转化体特异性PCR检测方法。结果显示：T-DNA插入片段全长4 318bp,由一个G2-EPSPS基因的表达盒构成。根据T-DNA 5'、3'端侧翼序列设计引物,建立了转化体特异性检测方法,并分析了该方法的灵敏度以及检出限,研究结果表明,3'端定性PCR检测方法特异性强,灵敏度高,检出极限为每100ng模板量的0.05%。本研究结果对转抗草甘膦外源基因检测和生物安全评价及监管具有重要意义。
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| 关 键 词： | 转基因玉米 G2-EPSPS 侧翼序列 转化体特异性检测 |
| 收稿时间： | 2015-10-20 |
Analysis of the Flanking Sequence and Event-Specific Detection of Transgenic G2-EPSPS Line of Maize |
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| Cui Guo,Wei Zhang,Guirong Yu,Zhengfu Zhou,Liang Li,Shuai Feng,Ming Chen,Jin Wang.Analysis of the Flanking Sequence and Event-Specific Detection of Transgenic G2-EPSPS Line of Maize[J].Crops,2016,32(1):69-20. |
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| Authors: | Cui Guo Wei Zhang Guirong Yu Zhengfu Zhou Liang Li Shuai Feng Ming Chen Jin Wang |
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| Institution: | 1 Life Science and Engineering College,Southwest University of Science and Technology,Mianyang 621010,Sichuan,China2 Biotechnology Research Institute,Chinese Academy of Agriculture Sciences,Beijing 100081,China3 Institute of Biological & Nuclear Technology,Sichuan Academy of Agricultural Sciences,Chengdu 610066,Sichuan,China; |
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| Abstract: | By hiTAIL-PCR(3'-terminal high-efficiency thermal asymmetric interlaced PCR) and LD-PCR(long distance PCR),the complete sequence and flanking sequence of T-DNA in transgenic maize D-3 were obtained. A specific PCR detection method for transformant was also established. The results revealed that the total length of T-DNA was 4318 bp,which consisted of a G2-EPSPS gene expression cassette. The primers had been designed based on the 3' and 5' terminal flanking sequence,followed by specific PCR to detect the sensitivity as well as the detection limit. The results showed that 3'-qualitative PCR detection method was highly specific and sensitive,with a sensitivity of 0.05% per 100ng DNA template.This research provides the methods for the detection of exogenous gene in transgenic maize and the risk assessment on bio-safety. |
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| Keywords: | Transgenic maize G2-EPSPS Flanking sequence Event-specific detection |
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