摘要
为获得进口转基因玉米筛查检测策略,并根据策略建立多重PCR方法,对15个进口转基因玉米分子特征进行分析。结果表明,将P-Ca MV 35S、T-NOS等2个基因组合作为检测策略可全部检出15个进口转基因玉米至少1次;将P-Ca MV 35S、P-ract1、T-NOS、bar、pat、PMI等6个基因组合作为检测策略,除MON810检出1次外,其他每个转化体可检出至少2次。同时,利用获得的筛查检测策略建立多重PCR体系,对2个基因组合的策略优化,结果表明适宜退火温度为58℃,适宜引物终浓度(μmol·L-1)配比为0.2∶0.2;对6个基因组合的策略优化,结果表明适宜退火温度为60℃,引物终浓度配比为0.2∶0.1∶0.1∶0.1∶0.1∶0.05。采用已知样品对多重PCR体系进行验证,图谱显示与转化体分子特征完全一致。该研究结果将为进口转基因玉米筛查检测提供参考。
In order to obtain screening test strategy on imported genetically modified （GM） maize and build the multiple polymerase chain reaction （PCR） methods based on the above strategy, molecular characteristics of 15 imported GM maize were analyzed. The results show that： 1） at least one time from 15 imported GM maize based on the testing strategy putting P-CaMV 35S and T-NOS together; 2） except for MON810, at least two times can be checked out according to the testing strategy contacting P-CaMV 35S, P-ract1, T-NOS, bar, pat with PMI. Meanwhile, the multiple polymerase chain reaction system was developed and optimized. The results show that the optimum annealing temperature of the dual PCR is 58 ℃, the optimum final concentration of primers（μmol·L^-1） ratio is 0.2：0.2, the optimum annealing temperature of the six-plex PCR is 60 ℃, the optimum final concentration of primers ratio is 0.2：0.1：0.1：0.1：0.1：0.05. Verification of the multiple PCR system with known samples the map showed that the amplified bands were identical with the molecular features of the GM maize. The results will provide valuable reference for the screening and testing strategy on imported GM maize.
出处
《核农学报》
CAS
CSCD
北大核心
2016年第6期1045-1053,共9页
Journal of Nuclear Agricultural Sciences
基金
四川省财政现代农业技术创新与示范专项资金项目(2014CXSF-040)
四川省教育厅自然科学一般项目(15ZB0331)
四川省科技厅项目(2014HH0031)
关键词
转基因玉米
多重PCR
筛查
检测
genetically modified maize, multiplex PCR, screening, detection
