Rapid Detection of CaMV-35S Promoter in Vegetable Oils by Loop - mediated Isothermal Amplification Method
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Abstract:
The Loop-mediated isothermal amplification (LAMP) assay is a novel and alternative DNA amplification method, which amplifies DNA with high specificity and efficiency, and has been successfully used to screen genetically modified (GM) crops (e.g., soybean, maize). In the present study, a LAMP assay for the detection of exogenous cauliflower mosaic virus 35 S promoter (CaMV-35S) in vegetable oils was firstly developed. The genomic DNA of vegetable oils was effectively extracted by adding hexane and co-precipitant. In the LAMP assay, and the reactions were optimized, involving different concentrations of FIP/BIP primer, betaine and Mg2+. Two different platforms of measurement (real-time turbidity and fluorescence dyes) were compared. Results indicated that LAMP method was rapid (within 56 min) and sensitive, and the detection sensitivity was ten fold higher than that of the conventional PCR. Without specialized PCR or electrophoresis instruments, the target DNA was amplified under isothermal conditions (63 ℃) and visualized by accumulated curve of turbidity or staining inspection (SYBR Green I) for naked-eye. The CaMV-35S promoter was successfully detected in vegetable oil samples (soybean oils and rapeseed oils) by this method. The LAMP approach was rapid, sensitive, and available for the monitoring of transgenic elements in GM vegetable oils.
