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Primer sequence and kit for real-time PCR detection of transgenic rape (seeds)

A primer sequence, rapeseed technology, applied in biochemical equipment and methods, microbial determination/inspection, etc., can solve the problems of complex process of genetically modified products, affecting the correct judgment of results, and many operation steps, so as to achieve rapid and sensitive detection technology. , the results are reliable, the effect of avoiding cross-contamination

Inactive Publication Date: 2007-04-11
曹际娟 +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The whole process of using this method to detect transgenic products is complicated, there are many steps, and post-PCR treatment is required, such as agarose gel electrophoresis and ethidium bromide staining, ultraviolet light observation results or detection by polyacrylamide gel electrophoresis or silver staining , not only need a variety of instruments, but also time-consuming and labor-intensive, the dye ethidium bromide used is harmful to the human body, and these complicated experimental processes provide opportunities for pollution and false positives, which seriously affect the correct judgment of the results

Method used

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  • Primer sequence and kit for real-time PCR detection of transgenic rape (seeds)
  • Primer sequence and kit for real-time PCR detection of transgenic rape (seeds)

Examples

Experimental program
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Effect test

Embodiment 1

[0015] The primer sequences and kits used for real-time PCR detection of SYBR(R) Green in transgenic rape (seed) RT73 strains are manufactured according to conventional primer preparation and synthesis methods to detect exogenous structural genes (transgenic components) of transgenic rape (seed) RT73 strains.

[0016] Detection gene: the foreign structural gene of RT73 strain.

[0017] Primers: 5’-cca tat tga cca tca tact ca ttg ct-3’, 5’-gct tat acg aag gca agaaaa gga-3’.

[0018] Kit: A kit for detecting the exogenous structural genes of the RT73 strain of transgenic rape (seed) using the primer sequence made by conventional methods.

Embodiment 2

[0020] The primer sequences and kits used for real-time PCR detection of SYBR(R) Green in the MS8 line of genetically modified rapeseed (seed) are manufactured according to conventional primer preparation and synthesis methods to detect exogenous structural genes (transgenic components) of the MS8 line of genetically modified rape (seed).

[0021] Detection gene: foreign structural gene of MS8 strain.

[0022] Primers: 5’-gac ctt tgc aat ttt ctc ttc agt act-3’, 5’-gcc ttt tct tat cga cca tgtact c-3’.

[0023] Kit: A kit for detecting exogenous structural genes of MS8 strains of transgenic rape (seed) made with the primer sequence.

Embodiment 3

[0025] The primer sequences and kits used in rape (seed) SYBR® Green real-time PCR detection are manufactured according to conventional primer preparation and synthesis methods to detect endogenous genes in transgenic rape (seed).

[0026] Detection gene: FatA gene.

[0027] Primers: 5’-ggt ctc tca gca agt ggg tga t-3’, 5’-tcg tcc cga act tca tct gta a-3’.

[0028] Kit: A kit for detecting endogenous FatA gene in rape (seed) made by using the primer sequence.

[0029] Main instruments for testing: real-time PCR instrument, ultra-clean workbench, sterilization pot, ice maker, nucleic acid protein analyzer, high-speed refrigerated centrifuge, desktop mini centrifuge, Mini personal centrifuge, low-temperature refrigerator, refrigerated freezer, Water purifier, double evaporator, vortex shaker, micro sampler (0.5μL, 2μL, 10μL, 20μL, 100μL, 200μL, 1000μL) etc.

[0030] Main reagents for detection: 10×PCR buffer, MgCl 2 , DNTP (dATP, dUTP, dCTP, dGTP), UNG enzyme (Uracil N-glycosylase),...

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Abstract

The present invention discloses the guide object sequence and reagent box used for real-time PCR inspection of transgenic rapeseed SYBR Green, providing the guide object sequence for the inspection of extraneous structure gene of transgenic rapeseed RT73 system, extraneous structure gene of MS8 system and endogenous gene of FatA, as well as the maxima, minima and optima selection sequence and the reagent box produced in accordance with guide object sequence. The present technology is used for qualitative and quantitative inspection of transgenic product and has the advantages of high resolution and avoiding cross pollution causing pseudo-positive reaction. SYBR Green real-time PCR technology employs complete closed-tube method, and PCR post treatment is not needed, avoiding cross pollution causing pseudo-positive reaction. SYBR Green dye is also very stable at the extreme temperature demanded by PCR reaction, which can efficiently distinguish specific product, nonspecific product and guide object dipolymer to compensate its bad specificity. The inspection technology is fast, sensitive, simple in operation, time saving and energy saving, accurate and reliable.

Description

(1) Technical field: [0001] The invention relates to gene amplification and detection. (2) Background technology: [0002] With the continuous commercialization of genetically modified products, the safety issue has been concerned by the world, and disputes continue to arise. In order to protect human health, eliminate consumer concerns, and facilitate international trade and commodity circulation, many countries represented by the European Union have formulated safety management regulations for such special products after a brief dispute. management. Many countries such as Europe and Japan have also formulated corresponding limit regulations. The Ministry of Agriculture of our country also promulgated the management measures for genetically modified products this year. However, in order for consumers to understand or accept genetically modified products, various laws and regulations can be implemented. In addition to providing relevant information and increasing transparency, th...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68
Inventor 曹际娟刘善斌赵昕
Owner 曹际娟
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