A calibrator plasmid for quantitative analysis of insect resistant maize (Yieldgard MON 810.

Real-time PCR (RT-PCR) is the preferred method for the quantification of genetically modified organisms
(GMOs) and implementation of labeling regulations. The precision, sensitivity, and reproducibility of RTPCR
data depend on the use of external calibrators. In this investigation, a dual target plasmid designated
pRSETMON-02 comprising of MON 810 maize event specific and endogenous zein gene sequences in 1:1
ratio in tandem was constructed and validated. Commutability of plasmid DNA (pDNA) and genomic DNA
(gDNA) calibrators for the quantification of MON 810 maize was assessed by employing a TaqMan RT-PCR
targeting the P-35S and zein gene. Higher PCR efficiencies, good linearity and lower relative standard
deviation (RSD) values were associated with pRSETMON-02 as opposed to gDNA calibrants. pDNA calibrants
exhibited better performance characteristic in terms of closeness to the expected value of
unknown samples than their genomic counterparts. Short term stability study of the pRSETMON-02
plasmid stored at different temperatures showed that pDNA is stable for 45 days at �20, and 4 �C. The
results demonstrated that the developed dual target plasmid pRSETMON-02 is fit for the intended use
of quantifying MON 810 maize and is a better alternative to conventional seed powder calibrants.