A rapid loop-mediated isothermal amplification method for detection of the modified GM cry1A gene in transgenic insect-resistant cotton and rice

Abstract

Among the commercial genetically modified (GM) crops, the insect-resistant GM crops are the major cultivars that cry gene is introduced into. A cry1Ab/1Ac GM fusion gene (GFM cry1A) and a GM truncated cry1Ac gene (cry1Ac-Mon) is the key foreign gene employed for construction of GM crops by China researchers and Monsanto Technology LLC respectively. Here these two genes are entitled “GM cry1A” gene and a rapid visual loop-mediated isothermal amplification (LAMP) assay method for detection of GM cry1A in transgenic insect-resistant crops was established. The LAMP assay was performed at an optimal temperature of 65 °C for 60 min in the presence of a set of four specific primers recognized six distinct sequences of the GM cry1A gene. The rough detection limit to the GM cry1A in samples is as low as 0.01% (a weight ratio of transgenic insect-resistant rice/cotton to non-transgenic rice/cotton). Comparatively, the sensitivity of this LAMP method is 10 times over that of the conventional PCR method. Fifteen cultivars/events and five Bt strains with or without cry1A gene were analyzed using the LAMP method as well as PCR method. The results demonstrate that this LAMP method shows a distinct specificity to the GM cry1A gene comparing with PCR analysis. Therefore, this LAMP method will be a potential effective tool for screening the GM cry1A gene in GM crops which are widely plant in China and other developing countries.