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Primer for detecting EPSPS gene of glyphosate-tolerant transgenic soybean and processed product
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A technology of genetically modified soybeans and glyphosate resistance, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc.
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Problems solved by technology
There have been many reports on methods for detecting transgenes. The currently established method for detecting glyphosate-resistant transgenic soybeans and processed products is mainly based on the detection of the endogenous gene Lectin, 35S promoter, NOS terminator and the target gene EPSPS. The operation is cumbersome and due to The genetic structure of soybean products is damaged after high temperature treatment, making it difficult to detect
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Embodiment 1
[0028] Follow the procedure below for testing:
[0029] (1) DNA extraction of transgenic soybean samples to be tested
[0030] A. Weigh 0.1g of soybeans, grind them, and transfer them to a 1.5mL centrifuge tube. Add 600 μL of preheated lysis buffer, mix gently, and keep warm in a water bath at 65°C for 10 minutes;
[0031] B. Add an equal volume of 600 μL of phenol / chloroform into the tube, mix well by inverting up and down, and extract for 2 minutes;
[0032] C. Centrifuge at 12000g for 5min, draw the supernatant into a new centrifuge tube, add the same volume of sedimentation solution as the supernatant, mix well, leave at room temperature for 10min, centrifuge at 12000g for 5min, remove the supernatant, and keep the precipitate;
[0033] D. Add 60 μL of RNase to the precipitate, place it at 37°C for 2 minutes, mix it well with a pipette tip, dissolve the precipitate at 37°C, add 300 μL of buffer solution after 5 minutes, and mix up and down 10 times;
[0034] E. Take out...
Embodiment 2
[0047] Follow the procedure below for testing:
[0048] (1) DNA extraction of the tofu sample to be tested
[0049] A. Weigh 0.1g of tofu, cut it into pieces, and transfer it to a 1.5mL centrifuge tube. Add 600 μL of preheated lysis buffer, mix gently, and keep warm in a water bath at 65°C for 10 minutes;
[0050] B. Add an equal volume of 600 μL of phenol / chloroform into the tube, mix well by inverting up and down, and extract for 2 minutes;
[0051] C. Centrifuge at 12000g for 5min, draw the supernatant into a new centrifuge tube, add the same volume of sedimentation solution as the supernatant, mix well, leave at room temperature for 10min, centrifuge at 12000g for 5min, remove the supernatant, and keep the precipitate;
[0052] D. Add 60 μL of RNase to the precipitate, place it at 37°C for 2 minutes, mix it well with a pipette tip, dissolve the precipitate at 37°C, add 300 μL of buffer solution after 5 minutes, and mix up and down 10 times;
[0053] E. Take out the spin...
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Abstract
The invention relates to a rapid screening and detection technology for glyphosate-tolerant transgenic soybean and processed products, in particular to primer groups for detecting the commonly used inserted gene EPSPS of glyphosate-tolerant transgenic plants. Aiming at the EPSPS gene and according to the conserved sequence of the EPSPS gene, the invention designs two groups of specific primers. By adopting the two groups of specific primers and the loop-mediated isothermal nucleic acid amplification technology, the EPSPS gene can be rapidly, sensitively and specifically detected, so the products containing glyphosate-tolerant transgenic plant ingredients are screened. The primer can be provided with other reagents in the form of reagent kits and is used for nucleic acid amplification reaction. The method has the advantages of simple and convenient operation and good repetitiveness.
Description
technical field [0001] The invention relates to a method for rapid screening and detection of glyphosate-resistant transgenic soybeans and processed products by using loop-mediated isothermal amplification technology, in particular to a rapid detection of integrated genes contained in glyphosate-resistant transgenic soybeans and processed products Primer sequences used in EPSPS. Background technique [0002] The rapid development of plantgenetic engineering has promoted the emergence of many transgenic plants, and their impact on human health and environmental safety has attracted worldwide attention. Long-term and effective monitoring of transgenic plants and their products scientifically and the establishment of a rapid and effective detection method system are of great significance to human health and sustainable agricultural development. Due to concerns about the safety of genetically modified crops and their products and issues such as international trade, many countr...
Claims
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