The Combination of Quantitative PCR and Western Blot Detecting CP4‐EPSPS Component in Roundup Ready Soy Plant Tissues and Commercial Soy‐Related Foodstuffs

Xiao, Xiao; Wu, Honghong; Zhou, Xin; Xu, Sheng; He, Jian; Shen, Wenbiao; Zhou, Guanghong; Huang, Ming

doi:10.1111/j.1750-3841.2012.02718.x

Cited by 14 publications

(12 citation statements)

References 17 publications

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“…To test its usefulness in analyzing GMOs, the transgenic soybeans called Roundup® Ready (RR) 1 and 2 developed by the Monsanto Corporation were analyzed by using this QuantStudio™ 3D Digital PCR System. Many studies have been conducted on the RR1 soybean, mainly using PCR-based methods to detect and/or quantify the transgene, e.g., [37]- [44]. Recently, detection and quantification of RR2 using digital PCR were also reported [34].…”

Section: Introductionmentioning

confidence: 99%

Application of Digital PCR in the Analysis of Transgenic Soybean Plants

Wan¹,

Li²,

Wu³

et al. 2016

ABB

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Detection and quantification of transgenes are important in analyzing genetically modified organisms (GMOs). Quantitative polymerase chain reaction (qPCR) is commonly utilized for such purposes. However, qPCR has certain limitations in detecting and quantifying transgenes in GMOs, such as the need of certified reference materials, a standard curve, and possible affection by inhibitors. Therefore, alternative and possibly better methods are needed. Recent advances in digital PCR technologies have promised to allow accurate quantification of nucleic acids and therefore provided another useful technique to analyze GMOs. Thermo Fisher Scientific has recently commercialized the Applied Biosystems QuantStudio 3D digital PCR system that can be used for a wide range of applications involving nucleic acids. It will be beneficial to the scientific community to show the applicability of this digital PCR system in detecting and quantifying transgenes in GMOs. In the present study, the transgenes present in the Roundup® Ready Soybean (RR1, event 40-3-2) and Roundup Ready Soybean 2 (RR2, event MON89788) developed by Monsanto Corporation were analyzed by using this digital PCR system. The qPCR analysis results were included for comparison. Using specifically designed TaqMan assays, as low as 1% of the RR1 or RR2 soybean material was reliably detected and quantified on the dPCR platform. Therefore, digital PCR is a sensitive and reliable method to analyze the RR transgenic soybeans, and should be another useful tool for analyzing other transgenic plants.

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Section: Introductionmentioning

confidence: 99%

Application of Digital PCR in the Analysis of Transgenic Soybean Plants

Wan¹,

Li²,

Wu³

et al. 2016

ABB

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“…CP4-EPSPS was derived from the soil bacterium Agrobacterium sp. Strain CP4 and the enzyme activity of CP4-EPSPS could not be inhibited by glyphosate due to weak binding affinity, resulting in increased tolerance to glyphosate in transgenic plants expressing this gene ( Franz et al, 1997 ; Guicheney et al, 2009 ; Xiao et al, 2012 ). GAT encodes an N-acetyltransferase for acetylation of glyphosate, which is the basis of a new mechanism of glyphosate tolerance in GM plants ( Castle et al, 2004 ; Siehl et al, 2005 , 2007 ).…”

Section: Discussionmentioning

confidence: 99%

Co-expression of G2-EPSPS and glyphosate acetyltransferase GAT genes conferring high tolerance to glyphosate in soybean

Guo

Hong

et al. 2015

Front. Plant Sci.

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Glyphosate is a widely used non-selective herbicide with broad spectrum of weed control around the world. At present, most of the commercial glyphosate tolerant soybeans utilize glyphosate tolerant gene CP4-EPSPS or glyphosate acetyltransferase gene GAT separately. In this study, both glyphosate tolerant gene G2-EPSPS and glyphosate degraded gene GAT were co-transferred into soybean and transgenic plants showed high tolerance to glyphosate. Molecular analysis including PCR, Sothern blot, qRT-PCR, and Western blot revealed that target genes have been integrated into genome and expressed effectively at both mRNA and protein levels. Furthermore, the glyphosate tolerance analysis showed that no typical symptom was observed when compared with a glyphosate tolerant line HJ06-698 derived from GR1 transgenic soybean even at fourfold labeled rate of Roundup. Chlorophyll and shikimic acid content analysis of transgenic plant also revealed that these two indexes were not significantly altered after glyphosate application. These results indicated that co-expression of G2-EPSPS and GAT conferred high tolerance to the herbicide glyphosate in soybean. Therefore, combination of tolerant and degraded genes provides a new strategy for developing glyphosate tolerant transgenic crops.

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“…The results indicated that high temperature and/or pressure lead to serious DNA degradation and significantly reduce the level of detectable DNA. Xiao et al (2012) used polymerase chain reaction (PCR) and Western blotting methods to evaluate the degradation of the CP4-EPSPS gene and protein in various bean foodstuffs, such as dried bean curd crust and deep-fried bean curd, and reported inconsistent results between the two detection techniques. Previous studies focused mainly on the degradation of exogenous genes in transgenic soybean, transgenic corn, and transgenic maize (Gryson, 2010), and little information is available on exogenous Bt genes or Bt proteins in transgenic rice.…”

Section: Introductionmentioning

confidence: 99%