Development of Polymerase Chain Reaction Methods to Detect Plant DNA in Animal Tissues

Klaften, Matthias; Whetsell, A. J.; Webster, John; Grewal, Rupinder; Fedyk, Eric R.; Einspanier, Ralf; Jennings, J. C.; Lirette, R. P.; Glenn, K. C.

doi:10.1021/bk-2004-0866.ch006

Cited by 6 publications

(3 citation statements)

References 7 publications

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“…Because of the potential for laboratory-derived contamination, in this study both the PCR analyses and also the extraction procedures were controlled by inclusion of extraction-negative and PCR-negative controls. The importance of stringently controlled experimental procedures during analyses of high copy number genes was described in a recent report (21). The authors showed that, working under standard laboratory conditions, PCR methods similar to those used in this study detected a rbcL gene fragment in 48% of the tested bovine tissue samples.…”

Section: Pcr Analysis Of Animal Samples For Rbcl Dna Fragmentsmentioning

confidence: 51%

Sensitive PCR Analysis of Animal Tissue Samples for Fragments of Endogenous and Transgenic Plant DNA

Nemeth¹,

Wurz²,

Artim³

et al. 2004

J. Agric. Food Chem.

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An optimized DNA extraction protocol for animal tissues coupled with sensitive PCR methods was used to determine whether trace levels of feed-derived DNA fragments, plant and/or transgenic, are detectable in animal tissue samples including dairy milk and samples of muscle (meat) from chickens, swine, and beef steers. Assays were developed to detect DNA fragments of both the high copy number chloroplast-encoded maize rubisco gene (rbcL) and single copy nuclear-encoded transgenic elements (p35S and a MON 810-specific gene fragment). The specificities of the two rbcL PCR assays and two transgenic DNA PCR assays were established by testing against a range of conventional plant species and genetically modified maize crops. The sensitivities of the two rbcL PCR assays (resulting in 173 and 500 bp amplicons) were similar, detecting as little as 0.08 and 0.02 genomic equivalents, respectively. The sensitivities of the p35S and MON 810 PCR assays were approximately 5 and 10 genomic equivalents for 123 bp and 149 bp amplicons, respectively, which were considerably less than the sensitivity of the rbcL assays in terms of plant cell equivalents, but approximately similar when the higher numbers of copies of the chloroplast genome per cell are taken into account. The 173 bp rbcL assay detected the target plant chloroplast DNA fragment in 5%, 15%, and 53% of the muscle samples from beef steers, broiler chickens, and swine, respectively, and in 86% of the milk samples from dairy cows. Reanalysis of new aliquots of 31 of the pork samples that were positive in the 173 bp rbcL PCR showed that 58% of these samples were reproducibly positive in this same PCR assay. The 500 bp rbcL assay detected DNA fragments in 43% of the swine muscle samples and 79% of the milk samples. By comparison, no statistically significant detections of transgenic DNA fragments by the p35S PCR assay occurred with any of these animal tissue samples.

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Section: Pcr Analysis Of Animal Samples For Rbcl Dna Fragmentsmentioning

confidence: 51%

Sensitive PCR Analysis of Animal Tissue Samples for Fragments of Endogenous and Transgenic Plant DNA

Nemeth¹,

Wurz²,

Artim³

et al. 2004

J. Agric. Food Chem.

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“…Only in conjunction with optimized sample selection and processing, contamination-free DNA extraction, and suitable real-time PCR methods can valid data interpretation be provided on the basis of such extremely sensitive PCR assays (Klaften et al 2004). Several reports are available investigating cross platform studies and suitable calibrator or reference materials and providing different recommendations for detecting GMOs by real-time PCR (Terry, Shanahan et al 2002;Taverniers et al 2004Taverniers et al , 2005 The absolute quantification of gene fragments by use of PCR will, however, remain a challenge that depends mainly on the technique employed, on the instrumental hardware and software used, and, finally, on the availability of highly calibrated standards for each genetic modification.…”

Section: Nucleic Acid Amplifikation Methods Using Pcrmentioning

confidence: 99%

DNA‐based Methods for Detection of Genetic Modifications

Einspanier¹

2006

Genetically Engineered Food

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“…[4][5][6][7] However, increasing demands for early diagnosis of diseases and understanding of fundamental biological processes involved in disease development and progression are pushing the need for sensitive detection of biomarkers, especially at low levels. 8,9 Until now, some immunoassay methods including fluorescence immunoassay, 10,11 radioimmunoassay, 12 enzyme-linked immunosorbent assay (ELISA), 13 mass spectrometric immunoassay, 14,15 electrophoretic immunoassay, 16 chemiluminescence immunoassay, 17 and immune polymerase chain reaction (PCR) assay 18 have been used for clinical serum sample measurements. However there is still a critical demand for simple, rapid, sensitive and low-cost detection techniques for the earlier and sensitive profiling of cancer biomarkers, especially in point-of-care applications.…”

Section: Introductionmentioning

confidence: 99%

Ultrasensitive electrochemiluminescence immunoassay for tumor marker detection using functionalized Ru-silica@nanoporous gold composite as labels

Zhang

et al. 2012

Analyst

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In this work, we reported a simple and sensitive sandwich-type electrochemiluminescence (ECL) immunosensor for carcinoembryonic antigen (CEA) on a gold nanoparticles (AuNPs) modified glassy carbon electrode (GCE). The Ru-silica (Ru(bpy)(3)(2+)-doped silica) capped nanoporous gold (NPG) (Ru-silica@NPG) composite was used as an excellent label with amplification techniques. The NPG was prepared with a simple dealloying strategy, by which silver was dissolved from silver/gold alloys in nitric acid. The primary antibody was immobilized on the AuNPs modified electrode through l-cysteine and glutaraldehyde, and then the antigen and the functionalized Ru-silica@NPG composite labeled secondary antibody were conjugated successively to form a sandwich-type immunocomplex through the specific interaction. The concentrations of CEA were obtained in the range from 1 pg mL(-1) to 10 ng mL(-1) with a detection limit of 0.8 pg mL(-1). The as-proposed ECL immunosensor has the advantages of high sensitivity, specificity and stability and could become a promising technique for tumor marker detection.

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Development of Polymerase Chain Reaction Methods to Detect Plant DNA in Animal Tissues

Cited by 6 publications

References 7 publications

Sensitive PCR Analysis of Animal Tissue Samples for Fragments of Endogenous and Transgenic Plant DNA

Sensitive PCR Analysis of Animal Tissue Samples for Fragments of Endogenous and Transgenic Plant DNA

DNA‐based Methods for Detection of Genetic Modifications

Ultrasensitive electrochemiluminescence immunoassay for tumor marker detection using functionalized Ru-silica@nanoporous gold composite as labels

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