Visual and Rapid Detection of Two Genetically Modified Soybean Events Using Loop-mediated Isothermal Amplification Method
To develop effective alternatives for detecting genetically modified organisms (GMOs), we reported one optimized visual loop-mediated isothermal amplification (LAMP) method for the detection of exogenous DNA targets from two GM soybean events in this study. This isothermal amplification can be performed within 40 min without polymerase chain reaction (PCR) equipment and the derived LAMP products can be directly observed by naked eye employing SybrGreen I dye instead of conventional gel electrophoresis analysis. The limits of detection of these established visual LAMP assays were about 4 copies of haploid soybean genomic DNA, and which were much higher than those of reported conventional PCR assays. Furthermore, the high specificity of LAMP assays was determined. All the results demonstrated that the developed visual LAMP assays are convenient, cost-efficient, and rapid for on spot detection of GMOs.
KeywordsLoop-mediated Isothermal Amplification Visual Detection Genetically Modified Soybean
This work was supported by the National Key Basic Research Program (2007CB109201, 2007FY230100), the National Transgenic Plant Special Fund (2008ZX08012-002, 003,005, and 2009ZX08012-002B), the National Natural Science Foundation of China (30725022, 30700499), the national high-tech project “863” (2006AA10Z443), and Shanghai Chenguang project (2008CG16).
- Holst-Jensen A, Rønning SB, Løvseth A, Berdal KG (2003) PCR technology for screening and quantification of genetically modified organisms (GMOs). Anal Bioanal Chem 375:985–993Google Scholar
- Imai M, Ninomiya A, Minekawa H, Notomi T, Ishizaki T, Van Tu P, Tien NT, Tashiro M, Odagiri T (2007) Rapid diagnosis of H5N1 avian influenza virus infection by newly developed influenza H5 hemagglutinin gene-specific loop-mediated isothermal amplification method. J Virol Methods 141:173–180CrossRefGoogle Scholar
- James C (2008) Global status of Commercialized Biotech/GM Crops: 2008. ISAAA Briefs 39Google Scholar
- Misawa Y, Yoshida A, Saito R, Yoshida H, Okuzumi K, Ito N, Okada M, Moriya K, Koike K (2007) Application of loop-mediated isothermal amplification technique to rapid and direct detection of methicillin-resistant Staphylococcus aureus (MRSA) in blood cultures. J Infect Chemother 13:134–140CrossRefGoogle Scholar
- Nagamine K, Watanabe K, Ohtsuka K, Hase T, Notomi T (2001) Loop-mediated isothermal amplification reaction using a nondenatured template. Clin Chem 47:1742–1743Google Scholar
- Rodríguez-Lázaro D, Lombard B, Smith H, Rzezutka A, D'Agostino M, Helmuth R, Schroeter A, Malorny B, Miko A, Guerra B, Davison J, Kobilinsky A, Hernández M, Bertheau Y, Cook N (2007) Trends in analytical methodology in food safety and quality: monitoring microorganisms and genetically modified organisms. Trends Food Sci Technol 18:306–319CrossRefGoogle Scholar
- Zel J, Mazzara M, Savini C, Cordeil S, Camloh M, Stebih D, Cankar K, Gruden J, Morisset D, Van den Eede G (2008) Method validation and quality management in the flexible scope of accreditation: an example of laboratories testing for genetically modified organisms. Food Anal Methods 1:61–72CrossRefGoogle Scholar