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Method for quickly detecting transgenic corn by using LAMP method

A kind of genetically modified corn, a rapid technology, applied in the direction of biochemical equipment and methods, microbial determination/inspection, etc., can solve the problem of less detection of genetically modified corn, and achieve the effect of high specificity and easy identification

Inactive Publication Date: 2011-03-16
BEIJING AGRO BIOTECH RES CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The LAMP method is widely used in the qualitative and quantitative detection of infectious diseases including DNA viruses, RNA viruses, bacteria and parasites, but there are few reports on transgenic detection, especially in the detection of transgenic corn.

Method used

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  • Method for quickly detecting transgenic corn by using LAMP method
  • Method for quickly detecting transgenic corn by using LAMP method
  • Method for quickly detecting transgenic corn by using LAMP method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] A method for rapid detection of transgenic maize, which uses loop-mediated isothermal amplification reaction (LAMP) to detect the glufosinate-resistant Bar gene in transgenic maize.

[0032] The steps included in the method are as follows:

[0033] A. Prepare the reaction system in a 1.5-2.0ml Eppendoff tube;

[0034] In the described reaction system, include:

[0035] Betaine 0.3-1.5mol / L,

[0036] dNTP 0.2-3.5mmol / L,

[0037] Mg 2+ 2-18mmol / L,

[0038] Inner primers, outer primers; the concentration ratio of the inner primers and outer primers is 1-8:1;

[0039] The concentration of the inner primer is 1.6-0.2 μmol / L, and the concentration of the outer primer is 0.2 μmol / L;

[0040] Large fragment of Bst DNA polymerase, 10×Bst polymerse buffer, DNA template, enough ddH 2 O;

[0041] According to the bar gene sequence with GenBank accession number EU048867.1, use LAMP special primer design software http: / / loopamp.eiken.co.jp / e / index.html to design primers and...

Embodiment 2

[0061] This example is an optimization of the composition of the reaction system in the method for rapid detection of transgenic corn in Example 1. The quality and source of each component in the reaction system are the same as those in Example 1.

[0062] Include in this reaction system:

[0063] Betaine 0.3-1.3mol / L,

[0064] dNTP 1-2.5mmol / L,

[0065] Mg 2+ 6-14mmol / L,

[0066] Outer primers, inner primers, the concentration ratio of the inner primers and outer primers is 1-8:1,

[0067] The concentration of the inner primer is 1.6-0.2 μmol / L, and the concentration of the outer primer is 0.2 μmol / L;

[0068] Large fragment of Bst DNA polymerase, 10×Bst polymerse buffer, DNA template, enough ddH 2 O;

[0069] Introduce Mg in this embodiment 2+ The method is: add MgSO to the reaction system 4 Mother liquor, make Mg in this reaction system 2+ The concentration is 6-14 mmol / L.

[0070] The reaction system is 25 μl, or 50 μl.

Embodiment 3

[0072] This example is a preferred solution based on Example 2, and the quality and source of each component in the reaction system are the same as those in Example 2. Taking the system with a volume of 25 μl as an example, the reaction system includes:

[0073] Betaine 0.5mol / L,

[0074] dNTP 1mmol / L,

[0075] Mg 2+ 8mmol / L,

[0076] Outer primers, inner primers, the concentration ratio of the inner primers and outer primers is 8:1,

[0077] The concentration of the inner primer is 1.6 μmol / L, and the amount of the outer primer is 0.2 μmol / L;

[0078] Introduce Mg in this embodiment 2+ The method is: add MgSO to the reaction system 4 Mother liquor, make Mg in this reaction system 2+ The concentration is 8 mmol / L;

[0079] 8U Bst DNA polymerase large fragment 1μl, 10×Bst polymerase buffer 2.5μl, DNA template (6.5μg / ml concentration) 1μl, enough ddH 2 O;

[0080] The volume of the reaction system may also be 50 μl.

[0081] The primers and their sources used in t...

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Abstract

The invention relates to the field of transgenic plant detection, in particular to a method for quickly detecting transgenic corn. The method detects glufosinate-resistant Bar genes in the transgenic corn by using loop-mediated isothermal amplification (LAMP). The method does not need special instruments (such as PCR instrument), and is quicker and more economic compared with a conventional laboratory method. The method is simple and convenient in identification, has extremely high specificity, and can judge whether a destination fragment is amplified only by observing with naked eyes or detecting the precipitation turbidity with an ultraviolet imaging system of a turbidity meter so as to judge whether a genome of a core plant sample for detecting contains the glufosinate-resistant Bar genes, and a gel electrophoresis method for observation is not needed.

Description

technical field [0001] The invention relates to the field of transgenic plant detection, in particular to a method for rapidly detecting transgenic corn, which utilizes a loop-mediated isothermal amplification reaction (LAMP) to detect the glufosinate-resistant Bar gene in the transgenic corn. Background technique [0002] As one of the most important genetically modified crops in the world, transgenic corn has been planted on a large scale since 1996, and countries all over the world have started planting it. In 2007, the number of countries planting genetically modified corn has grown to 15, in order of planting area For the United States, Canada, Philippines, Argentina, South Africa, Spain, Portugal, France, Czech Republic, Uruguay, Honduras, Germany, Slovakia, Romania, and Poland. As of 2007, the global planting area of ​​genetically modified crops reached 100 to 100 million hectares 2 , of which the planting area of ​​transgenic corn is 35.2 million hm 2 , accounting ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 吴忠义黄丛林张秀海陈金松罗昌程曦梁宏霞李春华
Owner BEIJING AGRO BIOTECH RES CENT
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