Quadruple PCR detection of herbicide-resistant transgenic rapeseed T45

ObjectiveA multiple PCR detection met hod for herbicide-resistant transgenic rapeseed T45 was estab-lished, in order to provide a valuable reference method for detecting herbicide-resistant transgenic rapeseed T45.MethodThe endogenous gene CruA, three exogenous genes P-CaMV 35S, T-CaMV 35S and pat of rapeseed were se-lected as targets for quadruple PCR detection system, their specific primers were referenced to China national standards or reported literature. Then the reaction conditions, including annealing temperature and primer concentration, were opti-mized, the specificity and sensitivity were tested, and this system was validated by the known sample. Based on these re-searches, the quadruple PCR system for detecting herbicide-resistant transgenic rapeseed T45 were established. ResultThe results showed that, the optimum annealing temperature of quadruple PCR was 58℃, the optimum ratio for final con-centration (μmol/L ) of primers was 0 . 1∶0 . 2∶0 . 2∶0 . 2 for CruA∶P-CaMV 35S∶T-CaMV 35S∶pat, the sensitivity of the method was 0.01 ng/μL, and the amplified products of all the samples were completely consistent with the expectant molecular characteristics. ConclusionThe established quadruple PCR method for detecting herbicide-resistant transgenic rapeseed T45 can detect simultaneously endogenous gene and multiple exogenous genes of herbicide-resistant transgenic rapeseed T45 in a reaction system, so which can used as another efficient method to detect transgenic rapeseed T45.