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Comparison of droplet digital PCR with quantitative real-time PCR for determination of zygosity in transgenic maize.

Research paper by Xiaoli X Xu, Cheng C Peng, Xiaofu X Wang, Xiaoyun X Chen, Qiang Q Wang, Junfeng J Xu

Indexed on: 16 Sep '16Published on: 16 Sep '16Published in: Transgenic Research



Abstract

This study evaluated the applicability of droplet digital PCR (ddPCR) as a tool for maize zygosity determination using quantitative real-time PCR (qPCR) as a reference technology. Quantitative real-time PCR is commonly used to determine transgene copy number or GMO zygosity characterization. However, its effectiveness is based on identical reaction efficiencies for the transgene and the endogenous reference gene. Additionally, a calibrator sample should be utilized for accuracy. Droplet digital PCR is a DNA molecule counting technique that directly counts the absolute number of target and reference DNA molecules in a sample, independent of assay efficiency or external calibrators. The zygosity of the transgene can be easily determined using the ratio of the quantity of the target gene to the reference single copy endogenous gene. In this study, both the qPCR and ddPCR methods were used to determine insect-resistant transgenic maize IE034 zygosity. Both methods performed well, but the ddPCR method was more convenient because of its absolute quantification property.