A real-time fluorescent quantitative PCR for detection of genetically modified maize line MIR604

doi:10.3969/j.issn.1004-1524.2017.11.01

›› 2017, Vol. 29 ›› Issue (11): 1769-1774.DOI: 10.3969/j.issn.1004-1524.2017.11.01

• Crop Science • Next Articles

A real-time fluorescent quantitative PCR for detection of genetically modified maize line MIR604

CHANG Lijuan, SONG Jun^*, ZHANG Fuli, LIU Wenjuan, TANG Chunyan, WANG Dong, YIN Quan

Analysis and Test Center, Sichuan Academy of Agricultural Sciences, Chengdu 610066, China

Received:2017-04-07 Online:2017-11-20 Published:2017-12-05

Abstract

Abstract: Genetically modified maize MIR604 is an insect resistant maize line developed by Syngenta in 2005, which was approved as a raw material for food and feed processing in 2008 in China. In order to accurately detect MIR604 and its processing products, event-specific quantitative PCR detection method was established in this study. The PCR primers and probes were designed specially according to the left boundary sequence of MIR604 and maize genome sequence. Finally, we used the method to detect 6 kinds of non-genetically modified crops, MIR604 and other non-target transgenic crops. The results showed that the other samples apart from MIR604 were not detected. The measured value of the sample(CRM) containing 1% MIR604 was 1.07%, which was close to the real value (1%). The sensitivity experiment results indicated the method could detect only 5 copies of MIR604 molecular fragments. Thus, the event-specific quantitative PCR detection method of genetically modified maize MIR604 has high specificity, accuracy and sensitivity, which can be used for quantitative detection of genetically modified maize MIR604.

Key words: genetically modified maize MIR604, real-time fluorescent quantitative PCR, specificity, accuracy, sensitivity

CLC Number:

S513

CHANG Lijuan, SONG Jun, ZHANG Fuli, LIU Wenjuan, TANG Chunyan, WANG Dong, YIN Quan. A real-time fluorescent quantitative PCR for detection of genetically modified maize line MIR604[J]. , 2017, 29(11): 1769-1774.

References

[1] JAMES C. Global status of commercialized biotech/GM crops 2012[C]// ISAAA Briefs brief 44. Ithaca: International Service for the acquisition of Agri-Biotech Applications, 2013.
[2] 袁美妗,邓日强,胡晓晖,等. 苏云金芽孢杆菌Cry1Aa和Cry3A 结构域编码区的融合[J].农业生物技术学报,2002,10(3):287-290.
YUAN M J, DENG R Q, HU X H, et al. Fusion of Bacinus thuringiensis toxin Cry 1Aa and Cry 3A domain encoding sequence[J]. Journal of agricultural Biotechnology , 2012,10(3):287-290.(in Chinese with English abstract)
[3] 抗虫玉米MIR604及其衍生品种定性PCR方法: 农业部1485号公告-16-2010 [S]. 北京:中国农业出版社,2015.
[4] Event-specific method for the quantification of maize line MIR604 using real-time PCR v. 1.01[S/OL].[2013-11-06]. https://publications.europa.eu/en/publication-detail/-/publication/52caee36-c915-4bcd-8b14-4ddabda68683
[5] 转基因产品检测核酸定量PCR检测方法: GB19495.5-2004 [S]. 北京:中国标准出版社,2007.
[6] SCHOLTENS I M J, KOK E J, HOUGS L, et al. Increased efficacy for in-house validation of real-time PCR GMO detection methods[J]. Analytical & Bioanalytical Chemistry , 2010, 396(6): 2213-2227.
[7] WU G, WU Y H, NIE S J, et al. Real-time PCR method for detection of the transgenic rice event TT51-1[J]. Food Chemistry , 2010, 119(1):417-422.
[8] 王渭霞,赖凤香,洪利英,等. 实时定量PCR检测转基因水稻科丰6号插入拷贝数和转基因含量[J]. 农业生物技术学报,2012,20(1):9-15.
WANG W X, LAI F X, HONG L Y, et al. Copy number estimation and quantitative analysis of transgenic rice Kefeng 6 through real-time PCR strategies[J]. Journal of Agricultural Biotechnology , 2012, 20(1): 9-15.(in Chinese with English abstract)
[9] LI X, SHEN K L, YANG L T, et al. Applicability of a novel reference molecule suitable for event-specific detections of maize NK603 based on both 50 and 30 flanking sequences[J]. Food Control , 2010, 21(6):927-934.
[10] 宋君,雷绍荣,刘勇,等. 转基因玉米MON863品系特异定量PCR方法的建立[J]. 湖北农业科学,2011,50(24):5250-5253.
SONG J, LEI S R, LIU Y, et al. Establishment of constructing specific quantitative PCR of genetically modified maize( Zea mays L.)event MON863[J]. Southwest China Journal of Agricultural Sciences , 2011,50(24):5250-5253. (in Chinese with English abstract)
[11] ZHANG F L, SONG J, NIU B, et al. An event-specific qualitative and real-time PCR detection of 98140 maize in mixed samples[J]. Food Control , 2015, 57:1-8.
[12] 张广远,孙红炜,李凡,等. 转基因玉米 MIR162 转化事件特异性检测方法及其标准化[J].作物学报,2013,39(7):1141-1147.
ZHANG G Y, SUN H W, LI F, et al. Event-specific PCR detection method of genetically modified maize MIR162 and its standardization[J]. Acta Agronomica Sinica , 2013, 39(7): 1141-1147. (in Chinese with English abstract)
[13] TIGST D, INDIRA R. Multiplex qualitative PCR assay for identification of genetically modified canola events and realtime event-specific PCR assay for quantification of the GT73 canola event[J]. Food Control , 2008, 19:893-897.
[14] MANO J, FURUI S, TAKASHIMA K, et al. Development and validation of event-specific quantitative PCR method for genetically modified maize MIR604[J]. Shokuhin Eiseigaku Zasshi.Journal of the Food Hygienic Society of Japan , 2012, 53(4): 166-171.
[15] KIM J H, KIM H Y. Event-specific detection methods for genetically modified maize MIR604 using real-time PCR[J]. Food Science and Biotechnology , 2009, 18(5): 1118-1123.
[16] Definition of minimum performance requirements for analytical methods of GMO testing[S/OL].[2015-10-20]http://gmo-crl.jrc.ec.europa.eu/doc/MPR%20Report%20Application%2020_10_2015.pdf
[17] 转基因植物及其产品成分检测抗虫水稻TT51-1及其衍生品种定量PCR方法: 农业部2122号公告-8-2014[S]. 北京:中国农业出版社,2015.

A real-time fluorescent quantitative PCR for detection of genetically modified maize line MIR604

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