A simplified and accurate detection of the genetically modified wheat MON71800 with one calibrator plasmid

Kim, Jaehwan; Park, Saet-Byul; Roh, Hyo-Jeong; Park, Sunghoon; Shin, Min-Ki; Moon, Gui Im; Hong, Jin-Hwan; Kim, Hae‐Yeong

doi:10.1016/j.foodchem.2014.12.052

Cited by 13 publications

(7 citation statements)

References 22 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…The performance of pCAN as a calibrator in real-time PCR analysis was evaluated using three mixed DNA samples and 10 practical canola samples. Since the discrepancy of PCR efficiency between using plasmid and genomic DNA templates might influence the ratio of exogenous to endogenous genome copy number, a conversion factor (Cf) is often used. ,− , In general, a Cf value is defined as the genome copy number ratio of the event-specific DNA to endogenous reference gene in a homogeneous genomic DNA sample using the plasmid DNA CRM as a calibrator and calculated using the formula . To determine the Cf values of pCAN, homogeneous genomic DNAs of each GM event were prepared at three concentrations (10.0, 5.0, 1.0 ng/μL).…”

Section: Materials and Methodsmentioning

confidence: 99%

See 1 more Smart Citation

One Novel Multiple-Target Plasmid Reference Molecule Targeting Eight Genetically Modified Canola Events for Genetically Modified Canola Detection

Wang

et al. 2017

J. Agric. Food Chem.

View full text Add to dashboard Cite

Multiple-target plasmid DNA reference materials have been generated and utilized as good substitutes of matrix-based reference materials in the analysis of genetically modified organisms (GMOs). Herein, we report the construction of one multiple-target plasmid reference molecule, pCAN, which harbors eight GM canola event-specific sequences (RF1, RF2, MS1, MS8, Topas 19/2, Oxy235, RT73, and T45) and a partial sequence of the canola endogenous reference gene PEP. The applicability of this plasmid reference material in qualitative and quantitative PCR assays of the eight GM canola events was evaluated, including the analysis of specificity, limit of detection (LOD), limit of quantification (LOQ), and performance of pCAN in the analysis of various canola samples, etc. The LODs are 15 copies for RF2, MS1, and RT73 assays using pCAN as the calibrator and 10 genome copies for the other events. The LOQ in each event-specific real-time PCR assay is 20 copies. In quantitative real-time PCR analysis, the PCR efficiencies of all event-specific and PEP assays are between 91% and 97%, and the squared regression coefficients (R) are all higher than 0.99. The quantification bias values varied from 0.47% to 20.68% with relative standard deviation (RSD) from 1.06% to 24.61% in the quantification of simulated samples. Furthermore, 10 practical canola samples sampled from imported shipments in the port of Shanghai, China, were analyzed employing pCAN as the calibrator, and the results were comparable with those assays using commercial certified materials as the calibrator. Concluding from these results, we believe that this newly developed pCAN plasmid is one good candidate for being a plasmid DNA reference material in the detection and quantification of the eight GM canola events in routine analysis.

show abstract

Section: Materials and Methodsmentioning

confidence: 99%

“…Kim et al. developed a simple plasmid reference for GM wheat MON71800 . Pi et al reported a plasmid reference material targeting five GM soybean events .…”

Section: Introductionmentioning

confidence: 99%

One Novel Multiple-Target Plasmid Reference Molecule Targeting Eight Genetically Modified Canola Events for Genetically Modified Canola Detection

Wang

et al. 2017

J. Agric. Food Chem.

View full text Add to dashboard Cite

show abstract

“…또한, 제초제저항성 LM 옥수수 DAS-40278-9 (aad-1 유전자 삽입)와 LM 캐놀라 MON88302 (cp4 epsps 유전자 삽 입) PCR 반응 결과, 유전자변형 옥수수와 캐놀라에 대해서 만 특이적으로 도입유전자가 검출되는 것을 확인하였다 ( Fig. 3E and F Kim et al 2008;Kim et al 2015;Kim et al 2009b;Lee et al 2007;Min et al 2004;Moon et al 2012;Wu et al 2009;Yoo et al 2013 …”

Section: A-d)unclassified

Development of detection methods for six approved LM crops in Korea

Seol

Choi

et al. 2017

J Plant Biotechnol

View full text Add to dashboard Cite

This is an Open-Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.Abstract Living modified crops are genetically modified living organisms and are widely used in biotechnical research and desired goods. As the reliance on LM products, concerns about safety of LMOs have been continuously increased in South Korea. We established the detection methods for unintentional released LMOs in environmental conditions. To detect six LM event genes of 1 canola, 1 maize and 4 soybeans, PCR conditions were based upon consideration of the Joint Research Centre information. Genomic DNAs were isolated from LM samples and PCR analysis were performed using each event-specific primer pair. Event-specific genes of all events were efficiently recognized by our methods. To investigate the insertion site of LM genes in each genome, we verified PCR product sequence by DNA sequencing. These results suggest that the LM event-specific gene amplification can be efficiently developed. In addition, our detection method is fit for monitoring and post-management of LM crops in the environment.

show abstract

“…The disadvantages of matrix-based CRMs in GMO quantitative analysis include the narrow dynamic range of quantification, multiple and complex preparation procedures, high cost, and difficulty in obtaining the homogeneous candidate [ 15 , 16 ]. Compared with matrix-based CRMs, plasmid-DNA-based CRMs are good substitutes in GMO analysis since they have a well-characterized transgene sequence and copy number, and they can be easily maintained in and produced from bacterium stocks [ 17 , 18 , 19 , 20 , 21 ]. In the detection of foodborne bacteria and viruses, plasmid-DNA-based CRMs are also widely used instead of inactivated bacteria and viruses [ 22 , 23 ].…”

Section: Introductionmentioning

confidence: 99%

Collaborative Ring Trial of the Applicability of a Reference Plasmid DNA Calibrant in the Quantitative Analysis of GM Maize Event MON810

Meng

Wang

Guo

et al. 2022

Foods

View full text Add to dashboard Cite

Certified reference materials (CRMs) is one of the critical requirements in a quantitative analytical method, such as in the quantification of genetically modified (GM) contents in food/feed products. Plasmid-DNA-based CRMs are becoming essential in GM content quantification. Herein, we report the construction of one plasmid DNA calibrant, pMON810, for the quantification of the GM maize event MON810 which is commercially planted and used for food/feeds worldwide, and the collaborative ring trial was used to validate its applicability. pMON10 was proven to have high specificity for the MON810 event. The limit of detection (LOD) and limit of quantification (LOQ) of real-time PCR assays of MON810 event and maize endogenous gene using pMON810 as calibrant was 2 copies/μL and 5 copies/μL, respectively. A total of eight laboratories participated in the ring trial and returned valid test results. Each sample was performed with three repeats and three parallels in each repeat. Statistical analysis of the ring trial results showed that pMON810 as a calibrant had high PCR efficiency (ranging from 0.885 to 1.008) and good linearity (ranging from 0.9933 to 0.9997) in MON810 and endogenous gene real-time PCR assays. The bias between the test values and true values ranged from 4.60 to 20.00% in the quantification of five blind samples. These results indicate that pMON810 is suitable for use as a calibrant for the quantification of MON810 events in routine lab analysis or to evaluate detection methods for MON810, as well as being used as a substitute for the matrix-based CRM of MON810.

show abstract

A simplified and accurate detection of the genetically modified wheat MON71800 with one calibrator plasmid

Cited by 13 publications

References 22 publications

One Novel Multiple-Target Plasmid Reference Molecule Targeting Eight Genetically Modified Canola Events for Genetically Modified Canola Detection

One Novel Multiple-Target Plasmid Reference Molecule Targeting Eight Genetically Modified Canola Events for Genetically Modified Canola Detection

Development of detection methods for six approved LM crops in Korea

Collaborative Ring Trial of the Applicability of a Reference Plasmid DNA Calibrant in the Quantitative Analysis of GM Maize Event MON810

Contact Info

Product

Resources

About