From its discovery in the 1980s, Polymerase chain reaction was further developed and is nowadays used as the foundation for the various PCR-based techniques used in molecular diagnosis across different species, and numerous types of samples. Real-Time PCR enables the user to monitor the amplification of a deoxyribonucleic acid (DNA) or complementary DNA (cDNA) target during the PCR run, in real-time, and not at the end, as it is the case in conventional PCR. The most frequent types of applications include gene expression analysis, gene silencing, variant analysis, and fusion temperature analysis. Given its vast field of application, a key question remains, and it is related to the controls (negative controls, positive controls, internal exogenous and endogenous controls) and their purpose in a Real-Time PCR experiment. In this paper, we set out to find how and when to use them, and which type of controls are suitable for certain experiment types, since the use of appropriate controls during Real-Time PCR experiments will reduce the effects of variables aside from the independent variable within the sample, therefore yielding accurate results, be it in research or diagnostic purposes.
Objective. Amflora potato, event EH92-527-1 was genetically modified to produce only the amylopectin component from starch composition. The presence of the transgenic potato in the food on Romanian market should be verified although the Amflora potato uses for industrial purposes and animal feed. The aim of this study was to detect the presence of the Amflora potato in the food or feed products.
Methods. For this purpose, five samples of potato tubers and four samples of chips were analyzed. DNA isolation was performed with E.Z.N.A. ® MicroElute Genomic DNA kit (Omega Bio-Tek, USA). For identification a potential presence of the recombinant DNA in the food samples it was used GMOIdent RT Event EH92-527-1 Potato kit (Eurofins GeneScan, Germany), that is a real-time PCR kit for qualitative event-specific detection of AmfloraTM EH92-527-1 potato.
Results. Amflora transgenic potato was not detected in any of analyzed samples.
Conclusions. The results of real-time PCR method confirm the absence of Amflora event EH92-527-1 in all potato tubers and chips analyzed samples.