Quantification of CP4-EPSPS in genetically modified Nicotiana tabacum leaves by LC-MS/MS with ¹⁸O-labeling

The CP4-EPSPS gene is widely used in herbicide-tolerant plants/crops all over the world. In this study, a method was developed by coupling liquid chromatography with high sensitivity to tandem mass spectrometry to quantify the amount of CP4-EPSPS expression in Nicotiana tabacum leaves. The quantification of protein was converted to measure the unique peptide of CP4-EPSPS protein. One peptide unique to CP4-EPSPS was synthesized and labeled with H₂ ¹⁸O to get ¹⁸O stable isotope labeled peptide. The peptide served as the internal standard. The validated method had good specificity and linearity. The intra-and inter-day precisions and accuracy for all samples were satisfactory. The results demonstrated that the novel method was sensitive and selective to quantify CP4-EPSPS in the crude extract without time-consuming pre-separation or the purification procedures.