摘要
以A2704-12、A5547-127、MON89788和GTS-40-3-2等4种商品化耐除草剂转基因大豆为材料,构建质粒标准分子用于解决转基因作物检测时标准物质缺乏现状。采用双酶切技术,将扩增的大豆内参基因和耐除草剂转基因大豆转化体特异性片段克隆至pMD18-T载体上。结果显示,质粒标准分子定性PCR特异性序列片段的LOD均为20copies/μL,定量检测体系Bias最大值≤9.89%,CV最大值≤5.96%,SD最大值≤0.06,实验得到的所有偏差值均在允许范围之内,构建的质粒标准分子适用于4种耐除草剂转基因大豆特异性检测。
To find a solution to the lack of calibrator during GMO detection,four Genetically Modified soybean events GTS-40-3-2,MON89788,A2704-12and A5547-127were used to construct a calibration plasmid which was developed by combining endogenous reference gene and herbicide resistance specific DNA sequence to the clone vector pMD18-T. The results showed LOD(a limit of detection)value of calibrator plasmid DNA was 20copies/μL by qualitative PCR detection.The max Bias value≤9.89%,max CV value≤5.96%,max SD value≤0.06in the performance of this realtime Q-PCR system.The deviations were reasonable,thus the calibrator plasmid was suitable for the specific detection of the four Genetically Modified soybean events.
出处
《中国农业大学学报》
CAS
CSCD
北大核心
2014年第3期49-54,共6页
Journal of China Agricultural University
基金
国家转基因生物新品种培育重大专项(2013ZX08004-002-002-004)
哈尔滨市科技局项目(2010RFQXN101)
关键词
大豆
转基因大豆
质粒标准分子
实时荧光定量PCR
soybean
Genetically Modified soybean
calibrator plasmid DNA
quantitative PCR(Q-PCR)
