Development Of ELISA Kit For Detection Of Glyphosate-Resistant Genetically Modified Soybean

Abstract

The goal of research was development of competitive ELISA kit for detection of genetically modified plants resistant to glyphosate. As the source for gene isolation we have used leaves of GM soybean RoundUp Ready, line 40-3-2 (Monsanto). Using polymerase chain reaction the gene encoding the enzyme 5-enolpiruvilshykimat-3-phosphate synthase (CP4 EPSPS) was isolated, which provides resistance to the herbicide glyphosate. Isolated gene has been cloned into plasmid vector pET24(a), creating genetic construct pET-CP4EPSPS. E.coli (BL 21(DE3)) has been transformed by genetic construct and over expressed after induction by 0.5 mM IPTG. Recombinant enzyme CP4 EPSPS with molecular weight 45 kD was purified as inclusion bodies and used as antigen for 96-well polystyrene plates immobilization. Monoclonal antibodies CP4 EPSPS mAb3 (Artron BioResearch Inc., Canada) was used for synthesis of horseradish peroxidase conjugate and receiving diagnostic reagent. We have shown that in the competitive ELISA we can detect at least 0.1% of GM plants resistant to glyphosate, on the example of soybean (line RR 40-3-2), moreover we can also detect rapeseed (line GT73) and sugar beet (line H7-1).