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Method for rapidly detecting genetically modified corn MON863

A technology of MON863 and transgenic corn, applied in the field of molecular biology, can solve the problems of expensive instruments and equipment, high detection cost and expense, and high technical conditions of transgenic products, and achieve the effect of simple operation and convenient identification

Inactive Publication Date: 2011-06-01
CENT LAB TIANJIN ACADEMY OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the high technical conditions required for the detection of genetically modified products, the equipment is relatively expensive, and the detection cost and expense are relatively high

Method used

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  • Method for rapidly detecting genetically modified corn MON863
  • Method for rapidly detecting genetically modified corn MON863
  • Method for rapidly detecting genetically modified corn MON863

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] (1) Reagent: Bst DNA polymerase large fragment produced by BioLabs (NEW ENGLAND) and 10 times ThermoPol Buffer solution; specific primer mixture; 4mol / L betaine solution; 0.2mol / L MgSO 4 solution.

[0055] (2) Amplification reaction system: the total volume of the amplification reaction is 25 μL, and its various components are: 10×ThermoPol Buffer 2.5 μL, 4mol / L betaine 6.25 μL, 0.2mol / L MgSO4 0.25 μL, mixed primer 1 μL , 10μmol / L dNTPs 3.5μL, 8000U / L Bst DNA polymerase large fragment 1μL, template DNA 1μL, make up to 25μL with sterilized deionized water, mix well and centrifuge (4000rpm, 5 seconds) on the machine.

[0056] (3) Amplification reaction procedure: proceed at 63° C. for 60 minutes, keep at 80° C. for 2 minutes, and store at 4° C.

[0057] (4) After the amplification reaction is over, take 15 μL of the system solution, add 1 μL of 1000×SYBR Green fluorescent dye directly into the amplification tube, shake and mix well, and observe the result with the naked ...

Embodiment 2

[0059] (1) Reagent: Bst DNA polymerase large fragment produced by BioLabs (NEW ENGLAND) and 10 times ThermoPol Buffer solution; specific primer mixture; 4mol / L betaine solution; 0.2mol / L MgSO 4 solution.

[0060] (2) Amplification reaction system: the total volume of the amplification reaction is 25 μL, and its various components are: 10×ThermoPol Buffer 2.5 μL, 4mol / L betaine 6.25 μL, 0.2mol / L MgSO 4 , mixed primers 0.25 μL, 10 μmol / L dNTPs 3.5 μL, 8000U / L Bst DNA polymerase large fragment 2 μL, template DNA 2 μL, made up to 25 μL with sterilized deionized water, mixed well, centrifuged (8000rpm, 10 seconds) machine.

[0061] (3) Amplification reaction procedure: proceed at 65°C for 45 minutes, keep at 80°C for 2 minutes, store at 4°C.

[0062] (4) After the amplification reaction is over, take 15 μL of the system solution, add 2 μL of 1000×SYBR Green fluorescent dye directly into the amplification tube, shake and mix well, and observe the result with the naked eye. The re...

Embodiment 3

[0064] (1) Reagent: Bst DNA polymerase large fragment produced by BioLabs (NEW ENGLAND) and 10 times ThermoPol Buffer solution; specific primer mixture; 4mol / L betaine solution; 0.2mol / L MgSO 4 solution.

[0065] (2) Amplification reaction system: the total volume of the amplification reaction is 25 μL, and its various components are: 10×ThermoPol Buffer 2.5 μL, 4mol / L betaine 6.25 μL, 0.2mol / L MgSO 4 , mixed primers 0.25 μL, 10 μmol / L dNTPs 3.5 μL, 8000U / L Bst DNA polymerase large fragment 2 μL, template DNA 5 μL, made up to 25 μL with sterilized deionized water, mixed well, centrifuged (5000rpm, 5 seconds) machine.

[0066] (3) Amplification reaction procedure: proceed at 63° C. for 60 minutes, keep at 80° C. for 2 minutes, and store at 4° C.

[0067] (4) After the amplification reaction was completed, 25 μL of the system solution was taken and analyzed by 2% agarose gel electrophoresis, and the results were observed under ultraviolet light. The reaction tubes without amp...

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Abstract

The invention discloses a method for rapidly detecting genetically modified corn MON863. The method adopts the principle as follows: four special primers and one DNA polymerase having chain replacemenThe invention discloses a method for rapidly detecting genetically modified corn MON863. The method adopts the principle as follows: four special primers and one DNA polymerase having chain replacement activity are adopted; and nucleic acid is augmented at 63-65 DEG C to be 10<9> to 10<10> copies in 45 to 60 min. An operator uses naked eyes to directly observe the turbidness of precipitation in at activity are adopted; and nucleic acid is augmented at 63-65 DEG C to be 10<9> to 10<10> copies in 45 to 60 min. An operator uses naked eyes to directly observe the turbidness of precipitation in areaction tube or judge whether the augmentation happens or not by the change in color after SYBR Green is added. The method can specially, rapidly, efficiently and conveniently detect the geneticallyreaction tube or judge whether the augmentation happens or not by the change in color after SYBR Green is added. The method can specially, rapidly, efficiently and conveniently detect the geneticallymodified corn MON863.modified corn MON863.

Description

technical field [0001] The invention belongs to the technical field of molecular biology, and relates to a detection method for transgenic products, specifically a method for rapidly detecting transgenic corn MON863, by visually observing the turbidity of a reaction tube or observing the color change after adding 1000×SYBR Green or observing the agarose Gel electrophoresis to judge the amplification. Background technique [0002] Genetically Modified Crops (GMC for short) refers to the use of modern genetic engineering technology to transfer certain biological genes into crops to modify the genetic material of crops so that they can improve their performance in traits (such as resistance, yield, etc.) , ripening period), quality and other aspects are changed to the goals required by human beings, and the crops obtained in this way are called "transgenic crops", also known as "genetically modified crops" or "genetically modified crops". [0003] Maize is one of the most impo...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12N15/11
Inventor 兰青阔王永程奕赵新朱珠
Owner CENT LAB TIANJIN ACADEMY OF AGRI SCI