The impact of analytical quality criteria and data evaluation on the quantification of genetically modified organisms

Meyer, William A.; Caprioara-Buda, M.; Jeynov, Boyan; Corbisier, Philippe; Trapmann, Stefanie; Emons, Hendrik

doi:10.1007/s00217-012-1787-7

Cited by 7 publications

(2 citation statements)

References 7 publications

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“…Recently, additional methods have been developed Consequently, in the current study, a qualitative PCR detection method for detecting the nine targets was developed. Highly linear and efficient reactions indicated that the standard curves of the pGMT27 plasmid were suitable for further quantitative test [32]. Meanwhile, using the pGMT27 plasmid as a calibrator provided a highly efficient method in the transgenic detection of flue-cured tobacco.…”

Section: Discussionmentioning

confidence: 99%

A New Reference Plasmid “pGMT27” Provides an Efficient Transgenic Detection Method for Flue-Cured Tobacco

Zhang

Adil

et al. 2021

Journal of Food Quality

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Owing to the economic value of its foliage, tobacco (Nicotiana tabacum) is cultivated all across the world. For the detection of genetically modified (GM) tobacco, there is a lack of universal standard material which ultimately limits the detection methods because the accuracy and comparability of the results cannot be ensured. Here, we prepared a reference plasmid “pGMT27” for the detection of GM tobacco, which was 18,296 bp in length harboring two of the tobacco endogenous and seven exogenous genes. By using qualitative PCR test for the nine genes, 10 copies were used for plasmid sensitivity. In the quantitative real-time PCR (qPCR) assays with pGMT27 as a calibrator, the reaction efficiencies for P-35S and NR were 101.427% and 98.036%, respectively, whereas the limit of detection (LOD) and limit of quantification (LOQ) were 5 copies and 10 copies per reaction. For standard deviation (SD) and relative standard deviation (RSD) of the Ct values, the repeatability values were from 0.04 to 0.42 and from 0.18% to 1.29%, respectively; and the reproducibility values were from 0.04 to 0.39 and from 0.18% to 1.14%, respectively. For the unknown sample test, the average conversion factor (Cf) was 0.39, and the accuracy bias was from −15.55% to 1.93%; for precision, the SD values ranged from 0.02 to 0.62, while RSD values were from 1.34% to 10.6%. We concluded that using the pGMT27 plasmid as a calibrator provided a highly efficient transgenic detection method for flue-cured tobacco.

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Section: Discussionmentioning

confidence: 99%

A New Reference Plasmid “pGMT27” Provides an Efficient Transgenic Detection Method for Flue-Cured Tobacco

Zhang

Adil

et al. 2021

Journal of Food Quality

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“…DNA degradation induced by processing was evaluated by employing qPCR assays that allowed simultaneous detection of RRS gene, as well as soybean-specific lectin gene in RRS construct-specific reaction. PCR efficiency and coefficient of determination (R 2 ) of qPCR calibration curves were identified as parameters of reliable determination of GMO content (32). Both standard curves showed a high degree of correlation (R 2 ≥0.998) in all the assays for the epsps and lectin genes, while PCR efficiencies ranged from 99 to 101 %, indicating the convenience of the standard curves for quantification.…”

Section: Quantitative Pcrmentioning

confidence: 99%

Untitled

2015

Food Technol. Biotechnol.

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SummaryThe aim of this study is to assess DNA degradation, DNA amplification, and GMO quantity during tofu production. Soybean seeds were spiked with Roundup Ready ® soybeans (RRS) at 0.9, 2, 3 and 5 % (by mass), to assess the level of RSS that would be of practical interest for threshold labelling. Real-time polymerase chain reaction (PCR) was more effective than conventional PCR in the analysis of raw soymilk, okara, boiled soymilk and tofu. The negative effect of grinding and mechanical manipulation was obvious in the okara sample prepared with 3 and 5 % RRS, where GMO content was reduced to (2.28±0.23) and (2.74±0.26) %, respectively. However, heating at 100 °C for 10 min did not cause significant degradation of DNA in all samples. The content of RRS in the final product, tofu, was reduced tenfold during processing, ranging from 0.07 to 0.46 %, which was below the labelling threshold level. The results are discussed in terms of global harmonization of GMO standards, which could have the positive effect on the trade of lightly processed foodstuffs such as tofu, especially regarding the labelling policies.

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GMO quantification: valuable experience and insights for the future

et al. 2014

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Cultivation and marketing of genetically modified organisms (GMOs) have been unevenly adopted worldwide. To facilitate international trade and to provide information to consumers, labelling requirements have been set up in many countries. Quantitative real-time polymerase chain reaction (qPCR) is currently the method of choice for detection, identification and quantification of GMOs. This has been critically assessed and the requirements for the method performance have been set. Nevertheless, there are challenges that should still be highlighted, such as measuring the quantity and quality of DNA, and determining the qPCR efficiency, possible sequence mismatches, characteristics of taxon-specific genes and appropriate units of measurement, as these remain potential sources of measurement uncertainty. To overcome these problems and to cope with the continuous increase in the number and variety of GMOs, new approaches are needed. Statistical strategies of quantification have already been proposed and expanded with the development of digital PCR. The first attempts have been made to use new generation sequencing also for quantitative purposes, although accurate quantification of the contents of GMOs using this technology is still a challenge for the future, and especially for mixed samples. New approaches are needed also for the quantification of stacks, and for potential quantification of organisms produced by new plant breeding techniques.

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The impact of analytical quality criteria and data evaluation on the quantification of genetically modified organisms

Cited by 7 publications

References 7 publications

A New Reference Plasmid “pGMT27” Provides an Efficient Transgenic Detection Method for Flue-Cured Tobacco

A New Reference Plasmid “pGMT27” Provides an Efficient Transgenic Detection Method for Flue-Cured Tobacco

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GMO quantification: valuable experience and insights for the future

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