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Development of a loop mediated isothermal amplification (LAMP) based detection platform for genetically modified organism (GMO) detection

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2019
Moğol, Ayça Nazlı
Genetically modified organisms (GMOs) are being widely used worldwide. Every country has a different legislation regarding the allowed events and GMO levels. This creates the great need for GMO detection. In this study, LAMP assay was used for GMO detection owing to its high sensitivity with the genetically modified organisms; Bt11 maize, GT73 Roundup Ready canola, and transgenic Nicotiana tabacum, targeting the sequences most commonly used in GMO constructions; 35S promoter and Figwort mosaic virus (FMV) sequences. Optimal conditions for the LAMP assay have been determined and with these conditions, sensitivity and specificity tests of the LAMP assays of 35S promoter were done using Bt11 maize and Nicotiana tabacum. The sensitivity and specificity tests of the LAMP assays of FMV were done using GT73 Roundup Ready canola. Hydroxy naphthol blue dye was used for monitoring of the LAMP products in addition to the agarose gel (1.5%) electrophoresis. The sensitivity tests showed that GMO detection was possible for as low as 1 double stranded genomic DNA copy for both 35S promoter and FMV sequences. The specificity tests showed that both of the primer sets (35S promoter and FMV) used in this study were highly specific and their specificity was not affected by the presence of a foreign DNA. Rapid DNA extraction techniques were also tested with the LAMP assay, and it was found that DNA extraction step could be finished as short as 5 minutes. Finally, lyophilized LAMP assay successfully detected 35S promoter sequence in Bt11 maize.