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Establishment of a Real-time Fluorescent Quantitative PCR Assay for Detection of genetically modified maize Line MON88017

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Agricultural Biotechnology 2017年第1期6卷 15-19,22页

作者： Jun SONG Dong WANG Analysis and Test Center Sichuan Academy of Agriculture SciencesChengdu 610066China

In order to improve the standardized technical systems of quantitative analyses for genetically modified organisms （GMOs） and products, ensure bio-safety and reduce ecological risk in China, a real-time fluorescent quantitative PCR assay was established for detection of genetically modified maize line MON88017. The established method was evaluated based on the specificity, sensitivity, accuracy and measurement uncertainty. The results showed that the established method had strong specificity in detection of genetically modified maize line MON88017. 1.50% MON88017 sample was detected with 29 replica- tions. The average measured value （ 1. 541% ） was close to the actual value （ 1.50% ） and the relative deviation was 2.70%. The variation coefficient of the measured value was 0.110 g ; the recovery was 100.00% and the measurement uncertainty was 0. 096. The limit of detection for genetically modified maize line MON88017 with the established method was 5 copies at the 97.5% confidence level. Thus, the real-time fluorescent quantitative PCR assay established in this study exhibited high specificity, accuracy and sensitivity, which could provide technical support for the safety supervision of genetically modified organ- isms and products in China.

关键词： genetically modified maize Real-time fluorescent quantitative PCR Specificity Sensitivity Accuracy Measurement uncertainty

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Development of an Event-specific Quantitative PCR for genetically modified maize (Zea mays) Event NK603

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Agricultural Biotechnology 2012年第5期1卷 20-23页

作者： Jun SONG Shaorong LEI Yong LIU Quan YIN Dong WANG Bing XIANG Fuli ZHANG Wenjuan LIU Lijuan CHANG Analysis and Test Center Sichuan Academy of Agricultural Sciences Institute of Plant Protection Sichuan Academy of Agricultural Sciences

[ Objective ] The aim of this study was to develop a quantitative PCR detection method for genetically modified maize event NK603, so as to provide ba- sis for quantitative analysis of event NK603. [ Methods ] A quantitative PCR detection method for genetically modified maize event NK603 was developed using primers and Taqman probe designed according to the flanking sequence of event NK603, which was then adopted to detect the samples containing 2% NK603 stand- ard （with uncertain quantity of 10% ）. [ Results ] The slope of standard curve ranged between -3.6 and -3.1, and the correlation coefficient was higher than 0. 99. The amplification efficiency of this method reached 100.2%, fallen between 90% and 110%. The detected quantity of the experimental sample was 1.9%, closer to the true quantity （2%）. [ Conclusion] This quantitative PCR detection method for genetically modified maize event NK603 is very precise and can be a- dopted in routine testing analysis.

关键词： genetically modified maize Event NK603 zSSIIb gene Flanking sequence Quantitative detection

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Establishment of a Construct-specific Real-time PCR Method for Quantitative Detection of genetically modified maize Line MIR604

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Agricultural Biotechnology 2016年第2期5卷 24-26页

作者： Lijuan CHANG Jun SONG Shaorong LEI Quan YIN Dong WANG Wenjuan LIU Fuli ZHANG Analysis and Test Center Sichuan Academy of Agriculture Sciences

In this study, a construct-specific real-time PCR method for quantitative detection of genetically modified maize line MIR604 was established with prim- ers and probes designed based on vector sequence of MIR604 under optimized reaction system and thermal cycling condition. By using the established method, six non-genetically modified crops, genetically modified maize line MIR604 and other non-target genetically modified crops were detected. According to the results, flu- orescence signal could be detected in genomie DNA of MIR604, but other non-genetically modified crops and non-target genetically modified crops exhibited no fluo- rescence signal of MIR604 molecular fragment. Certified reference materials containing 1% MIR604 were detected with the established method and the results indi- cated that the average relative content in the test samples was 1.05%. The sensitivity analysis results indicated that MIR604 nucleic acid fragment with at least five copies could be detected with the established method. In conclusion, the construct-specific real-time PCR method for quantitative detection of genetically modified maize line MIR604 is of high specificity, accuracy and sensitivity, which provides technical support for safety supervision of genetically modified organisms in China.

关键词： genetically modified maize Construct-specificity Sensitivity Accuracy

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Expression profiles of Cry1Ab protein and its insecticidal efficacy against the invasive fall armyworm for Chinese domestic GM maize DBN9936

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Journal of Integrative Agriculture 2021年第3期20卷 792-803页

作者： LIANG Jin-gang ZHANG Dan-dan LI Dong-yang ZHAO Sheng-yuan WANG Chen-yao XIAO Yu-tao XU Dong YANG Yi-zhong LI Guo-ping WANG Li-li GAO Yu YANG Xue-qing YUAN Hai-bin LIU Jian ZHANG Xiu-jie WU Kong-ming Development Center of Science and Technology Ministry of Agriculture and Rural AffairsBeijing 100176P.R.China State Key Laboratory for Biology of Plant Diseases and Insect Pests Institute of Plant ProtectionChinese Academy of Agricultural SciencesBeijing 100193P.R.China State Key Laboratory of Cotton Biology Institute of Cotton ResearchChinese Academy of Agricultural SciencesAnyang 455000P.R.China Shenzhen Branch Guangdong Laboratory for Lingnan Modern AgricultureGenome Analysis Laboratory of the Ministry of AgricultureAgricultural Genomics Institute at ShenzhenChinese Academy of Agricultural SciencesShenzhen 518120P.R.China Institute of Plant Protection and Soil Science Hubei Academy of Agricultural SciencesWuhan 430064P.R.China College of Horticulture and Plant Protection Yangzhou UniversityYangzhou 225009P.R.China Institute of Plant Protection Henan Academy of Agricultural SciencesZhengzhou 450002P.R.China Yantai Academy of Agricultural Sciences Yantai 265500P.R.China College of Plant Protection Shenyang Agricultural UniversityShenyang 110866P.R.China College of Plant Protection Jilin Agricultural UniversityChangchun 130118P.R.China College of Agriculture Northeast Agricultural UniversityHarbin 150030P.R.China

The fall armyworm(FAW)Spodoptera frugiperda,which originated in the Americas,is advancing across China and threatening the nation’s maize ***,one widely used tool for its control is genetically modified(GM)Bacillus thuringiensis(Bt)*** content of Bt protein in appropriate plant parts is crucial for enhancing resistance against insect *** this study,we conducted a systematic investigation of Cry1 Ab levels in Chinese domestic GM maize DBN9936,which has recently obtained a biosafety certificate,and evaluated its efficacy against *** of expression levels of Cry1 Ab,via ELISA,indicated a spatio-temporal dynamic,with significant variation of mean Cry1 Ab,ranging from 0.76 to 8.48μg g-1 FW with the Cry1 Ab protein level ranked as:V6-V8 leaf>R1 leaf>R4 leaf>R1 silk>VT tassel>R4 *** the nine locations,the Cry1 Ab levels in DBN9936 of the Xinxiang,Langfang,and Harbin fields were significantly lower than those from Wuhan and Shenyang,and were slightly,but not significantly lower than those from the other four ***,the artificial diet-Cry1 Ab mixture and plant tissue feeding bioassays revealed that DBN9936 has high efficacy against *** insecticidal efficacy of different tissues against FAW larvae reached 34-100%with a descending order of lethality as follows:VT leaf>R4 leaf>R1 husk>R1 silk>VT tassel>R4 *** together,our results showed that Bt-Cry1 Ab maize DBN9936 has potential as a promising strategy to manage FAW.

关键词： fall armyworm genetically modified maize DBN9936 Cry1Ab expression control efficacy

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