This paper presents the results of molecular studies on testing and evaluation of multiplex test systems for the genetically modified organisms’ (GMOs) identification by PCR in real time. For the detection of genetic modifications in plants of the cabbage family (Brassicaceae) screening of major regulatory sequences is insufficient, as in genetic engineering manipulations 35S promoter – of DNA-containing cauliflower mosaic virus (CaMV) is used, which presence in the test material can lead to false positive results. In this work the feasibility of testing for the presence of viral CaMV’s DNA and analysis of screened gene sequences of interest for the usage during the analysis of rapeseed (Brassica napus L.) samples. It was established that the analysis of the presence/absence of a positive result for the CaMV 35S promoter is not giving complete answers to the absence of GMO in the sample. The appropriateness of GMO’s screening in rape seeds with NOS terminator and genes of interest.

This paper presents the results of molecular studies on testing and evaluation of multiplex test systems for the genetically modified organisms’ (GMOs) identification by PCR in real time. For the detection of genetic modifications in plants of the cabbage family (Brassicaceae) screening of major regulatory sequences is insufficient, as in genetic engineering manipulations 35S promoter – of DNA-containing cauliflower mosaic virus (CaMV) is used, which presence in the test material can lead to false positive results. In this work the feasibility of testing for the presence of viral CaMV’s DNA and analysis of screened gene sequences of interest for the usage during the analysis of rapeseed (Brassica napus L.) samples. It was established that the analysis of the presence/absence of a positive result for the CaMV 35S promoter is not giving complete answers to the absence of GMO in the sample. The appropriateness of GMO’s screening in rape seeds with NOS terminator and genes of interest.

Purpose. To create a multiplex system for identification glyphosate-tolerant sugar beet by using PCR. Methods. Molecular genetic analysis. Results. The article presents the results of studies to determine the parameters of the polymerase chain reaction (PCR) in order to develop a multiplex system for identification of the structural elements of the design of transgenic gene cp 4 epsps, which provides tole­rance to glyphosate. For amplicon target DNA sequences, the following values of temperature conditions of PCR were determined: step 1 (initial denaturation) 95 °C – 3 min; step 2 (specific reaction products accumulation): denaturation 95 °C – 45 s; hybridization of primers 55 °C – 50 s; elongation 72 °C – 1 min; number of cycles – 40; step 3 (final elongation) 72 °C – 6 min. A series of PCR were carried out for the purpose of selecting the optimal amount of DNA matrix for efficient estimate of transgenic sugar beet plants for the presence of specific sequences. Conclusions. To identify transgenic glyphosate-tolerant sugar beet, it is advisable to determine 35S promoter and gene cp 4 epsps in individual genotypes. It was found that during the selection of temperature parameters of multiplex reaction a 5 °C rise in primer hybridization temperature did not affect the identification of gene als that allowed to include specific primers for determination of this sequence as an internal control. Based on the results of test multiplex reactions, concentrations of dNTPs and Mg2+ ions were determined that allowed to exclude the possibility of non-specific fragments and false-negative results. The optimum amount of matrix DNA (100–150 ng) for an efficient estimate of transgenic sugar beet plants for the presence of specific sequences was determined. Obtained results allowed to develop a multiplex test system for identification of transgenic glyphosate-to­lerant sugar beet which can be used for simultaneous determination of the 35S promoter, cp 4 epsps gene and als gene as an internal reaction control.

Purpose. To create a multiplex system for identification glyphosate-tolerant sugar beet by using PCR. Methods. Molecular genetic analysis. Results. The article presents the results of studies to determine the parameters of the polymerase chain reaction (PCR) in order to develop a multiplex system for identification of the structural elements of the design of transgenic gene cp 4 epsps, which provides tole­rance to glyphosate. For amplicon target DNA sequences, the following values of temperature conditions of PCR were determined: step 1 (initial denaturation) 95 °C – 3 min; step 2 (specific reaction products accumulation): denaturation 95 °C – 45 s; hybridization of primers 55 °C – 50 s; elongation 72 °C – 1 min; number of cycles – 40; step 3 (final elongation) 72 °C – 6 min. A series of PCR were carried out for the purpose of selecting the optimal amount of DNA matrix for efficient estimate of transgenic sugar beet plants for the presence of specific sequences. Conclusions. To identify transgenic glyphosate-tolerant sugar beet, it is advisable to determine 35S promoter and gene cp 4 epsps in individual genotypes. It was found that during the selection of temperature parameters of multiplex reaction a 5 °C rise in primer hybridization temperature did not affect the identification of gene als that allowed to include specific primers for determination of this sequence as an internal control. Based on the results of test multiplex reactions, concentrations of dNTPs and Mg2+ ions were determined that allowed to exclude the possibility of non-specific fragments and false-negative results. The optimum amount of matrix DNA (100–150 ng) for an efficient estimate of transgenic sugar beet plants for the presence of specific sequences was determined. Obtained results allowed to develop a multiplex test system for identification of transgenic glyphosate-to­lerant sugar beet which can be used for simultaneous determination of the 35S promoter, cp 4 epsps gene and als gene as an internal reaction control.