Effect of the Matrix and Target on the Accurate Quantification of Genomic and Plasmid DNA by Digital Polymerase Chain Reaction

In polymerase chain reaction (PCR)-based nucleic acid quantification, the DNA template type, primer/probe sequence, and instrument platform such as real-time quantitative PCR (qPCR) digital (dPCR) affect accuracy reliability of results. this study, a plasmid (pDNA) pBI121-screening, genetically modified (GM) rice SDrice genomic (gDNA), GM rapeseed SDrape gDNA, all carrying same 11 screening elements, were used to prepare samples different levels gDNA pDNA in non-GM background. The comparison dPCR assays targeting elements revealed that set is key factor affects quantification. optimal method for was screened out from among validated qPCR methods. quantification low-level test low when calibrator, whereas high not affected by variations type detection target. one or two can be randomly selected characterize multiple-target reference materials (RMs). Low-level RMs with certified values quality controls avoid significant bias