Website: https://vjfc.nifc.gov.vn/
Website: https://vjfc.nifc.gov.vn/
The nopaline synthase terminator (T-nos) is a transcriptional termination sequence originally derived from the Ti (tumor-inducing) plasmid of Agrobacterium tumefaciens, a well-known soil bacterium capable of inducing crown gall disease in plants. In plant biotechnology, T-nos is widely employed as a functional regulatory element in expression cassettes to control the transcriptional termination of transgenes in genetically modified (GM) crops. The inclusion of T-nos ensures accurate termination of transcription, leading to the generation of stable and efficiently translatable mRNA transcripts, thereby enhancing the expression of the target gene. Due to its high compatibility with plant expression systems and consistent performance across diverse plant species, T-nos has become a standard terminator in the design of plant transformation vectors. The most frequently used method for detecting the presence of genetically modified material in food products is the screening of specific deoxyribonucleic acid (DNA) sequences, notably the CaMV 35S promoter and the T-nos terminator, using polymerase chain reaction (PCR) techniques. This study aimed to develop a real-time PCR assay targeting an 84 bp fragment of the T-nos sequence for the detection of GM material in milk-based matrices, using a TaqMan probe-based approach. Certified reference materials (CRMs) containing Roundup Ready soybean (GTS 40-3-2) and NK603 events were utilized for method development and optimization. The proposed method demonstrated high sensitivity, with a limit of detection (LOD) of 0.05%, and achieved 100% specificity and accuracy. These results indicate that the method is highly reliable and effective for the detection of trace amounts of GM-derived DNA containing the T-nos sequence in complex food matrices such as milk products.
real-time PCR, Nopaline synthase terminator, T-nos, GM.
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