Study On Detecting Method For Specificity Of Transgenic Soybean OsDREB3

Safety and impaci on the ecological environment of genetically modified foods has been the focus of attention. Most of the commercialized of genetically modified soybeans is glyphosate-resistant soybean, stress resistance genes due to the geographical environment and the global commercialization of the increase in demand for the commercialization are getting higher and higher, but so far there has not been established the detection techniques resilience gene of genetically modified soybeans in China.In this study, we set up the detecting techniques for specificity of transgenic soybean OsDREB3due to the51end and3’end flanking sequence information of genetically modified soybean OsDREB3, which had entered the environmental releasing or production test or got the security certification of environmental safety evaluation for genetically modified plants. The5’end flanking sequence of344bp covering the transformation vectors and transgenic soybean gene group was gained by chromosome walking technique, in the soybean genome on chromosome1, position for49,828,108-49,836,357;3’ end flanking sequence of a total length2189bp, the case of unknown gene sequences.The experiment was based on the5’ end flanking sequence information, specific qualitative and quantitative detection of genetically modified soybeans OsDREB3strains, event-specific detection of qualitative and quantitative cited material by amplified fragment covering soybean genome and the transformation of vector sequence information, in the detection process randomly select the four kinds of genetically modified soybean, were randomly selected two kinds of genetically modified com, two kinds of transgenic rice at the same time, event-specific qualitative and quantitative detection were carried out results showing that in addition to transgenic soybeans OsDREB3in the amplified fragment or outside of the amplification curve, the rest of genetically modified crops have not amplified fragments or amplification curve, indicating that the qualitative and quantitative detection methods to comply with the requirements of the event-specific detection. Qualitative detection of amplified fragment size of160bp. The detection sensitivity of0.1%than the EU detection limit (0.9%) and good reproducibility can be used to genetically modified soybeans OsDREB3qualitative detection. The design of quantitative PCR primers, amplification curve detection limit up to0.01%.multiplex PCR assay for genetically modified soybeans and transgenic soybean OsDREB3nested PCR method were set up. In this study, to select the transfected gene soybean GTS40-3-2, genetically modified soybean A2704-12and transgenic soybean OsDREB3DNA as to be detected gene template in multiplex PCR components were adjusted several times, a variety of different primers were tried on the ratio The amplified fragment size difference of more than100bases, ranging from700bp to200bp between good visibility. The best detection scheme is finalized, the successful implementation of three kinds of genetically modified soybeans, amplification and detection of four different fragments, The low limits of detection was0.01%. Nested PCR primers to meet the event-specific qualitative detection, but also to improve the detection sensitivity, this experiment created by the nested PCR method, and the sensitivity of the detection was0.005%.In this study, we constructed a standard molecular containing five kinds of transgenic soybean lines and soybean internal reference gene lection as the positive standard control. In this study, by real-time quantitative PCR, the soybean lectin gene (the lectin) as the endogenous reference gene to determine the crop exogenous gene OsDREB3in the genetically modified soybean OsDREB3genome. We found the OsDDREB3in soybean OsDreb3was only one copy.In this experiment, we built up a set of sensitive and high, with good reproducibility, high reliability, specificity and event-specific detection system of genetically modified soybean OsDREB3, by which we can improve detection efficiency and perfect insufficient of detection in modified soybean OsDREB3, and provide technical support for the detection of the the transgenic soybean OsDREB3, the results of this study provide a valuable reference for GMO labeling system specification.