A Multiplex, Fluorescent, and Isothermal Method for Detecting Genetically Modified Maize

Detecting, monitoring, and mapping transgenic sequences in the environment has been a long-standing challenge in ecology. Given the level of social, economic, and scientific interest in them, transgenic (GM) DNA sequences stand as paradigmatic; such sequences further have the unique property of functioning as unequivocal markers in the environment, clearly showing the paths of gene flow between crops and their wild relatives, and across industrial and traditional forms of agriculture. True mapping of transgenic DNA at the landscape level requires a method of DNA amplification that is robust and isothermal in order to be cost effective and field-based. We present here a method with these characteristics. Our multiplex fluorescent method is for the detection of the p35s CaMV promoter frequently used in transgenic plants as well as the alpha-zein protein specific to maize, but can be applicable to any other DNA sequence. The method uses loop-mediated isothermal amplification (LAMP) with visualization achieved using a fluorophore-quencher system of DNA hybridizing probes. We demonstrate the applicability of this tri-color method to transgenic corn, non-transgenic corn, and non-transgenic/non-corn species.