Extracting agent for extracting DNA from insect materials and application thereof

An extraction agent, insect technology, applied in DNA preparation, recombinant DNA technology and other directions, can solve the problems of unstable extraction results, need more samples, and take a long time, so as to improve the practical application range, simplify the extraction process, and simplify the operation. Effect

Active Publication Date: 2019-02-22
INST OF PLANT PROTECTION CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There are many traditional DNA methods, such as the traditional classic phenol-chloroform method, etc. The DNA obtained by these methods is of high purity and can meet the requirements of various tests, but the operation is cumbersome and time-consuming, and the reagents used have certain toxicity
For example, the salt extraction method, the yield is relatively low, and the purity is poor
Such as silica gel membrane ads...

Method used

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  • Extracting agent for extracting DNA from insect materials and application thereof
  • Extracting agent for extracting DNA from insect materials and application thereof
  • Extracting agent for extracting DNA from insect materials and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0064] Example 1. The extractant of the present invention and its application

[0065] The extractant of this embodiment is a buffer composed of the following components:

[0066]

[0067] The pH of the buffer is 8.0, and the solvent is distilled water.

[0068] Weigh 0.3028g Tris-base and 0.8775g NaCl, dissolve it in 80mL distilled water, adjust the pH to 8.0 with concentrated hydrochloric acid, add 2g polyvinylpyrrolidone K 40 (PVP K-40), 1mL Tween-20, and finally add distilled water to make the volume constant To 100mL.

[0069] Using the extracting agent of the present invention to extract DNA from a sample of Myzus persicae, the specific method is as follows:

[0070] 1. Put a single Myzus persicae (single, average weight 0.248mg) into a 1.5ml centrifuge tube, add 30μL of extract, and grind for 5s;

[0071] 2. Vortex for 3 seconds and quickly centrifuge for 2 seconds to remove impurities.

[0072] 3. Take 20 μL of the supernatant as a DNA sample and place it in a refrigerator at 4 d...

Embodiment 2

[0073] Example 2. The extractant of the present invention and its application (Buffer No. 1 in Table 1 of Orthogonal Design)

[0074] The extractant of this example is a buffer solution (No. 1 in Table 1) composed of the following components:

[0075]

[0076] The pH of the buffer is 8.0, and the solvent is distilled water.

[0077] The preparation method of the extractant is:

[0078] Weigh 0.3028g Tris-base and 0.2925g NaCl, dissolve it in 80mL distilled water, adjust the pH to 8.0 with concentrated hydrochloric acid, add 1g polyvinylpyrrolidone K 40 (PVP K-40), 1mL Tween-20, and finally add distilled water to make the volume constant To 100mL.

[0079] Using the extracting agent of the present invention to extract DNA from a sample of Myzus persicae, the specific method is as follows:

[0080] 1. Put a single Myzus persicae (single, average weight 0.248mg) into a 1.5ml centrifuge tube, add 30μL of extract, and grind for 5s;

[0081] 2. Vortex for 3s and centrifuge quickly for 2s.

[00...

Embodiment 3

[0083] Example 3. The extractant of the present invention and its application (Buffer No. 5 in Table 1 of Orthogonal Design)

[0084] The extractant in this example is a buffer solution (No. 5 in Table 1) composed of the following components:

[0085]

[0086] The pH of the buffer is 8.0, and the solvent is distilled water.

[0087] The preparation method of the extractant is:

[0088] Weigh 0.6057g Tris-base, 0.5850g NaCl, dissolve it in 80mL distilled water, adjust the pH to 8.0 with concentrated hydrochloric acid, add 4g polyvinylpyrrolidone K 40 (PVP K-40), 1mL Tween-20, and finally add distilled water to make the volume constant To 100mL.

[0089] Using the extracting agent of the present invention to extract DNA from a sample of Myzus persicae, the specific method is as follows:

[0090] 1. Put a single Myzus persicae (single, average weight 0.248mg) into a 1.5ml centrifuge tube, add 30μL of extract, and grind for 5s;

[0091] 2. Vortex for 3s and centrifuge quickly for 2s.

[0092]...

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PUM

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Abstract

The invention discloses an extracting agent for DNA extraction and application thereof. The extracting agent for DNA extraction is composed of Tris-base, NaCl, PVP and Tween-20, and can rapidly extract a trace amount of DNA from insect materials, and only the extracting agent, instead of other liquid, needs to be disposed in the sample extraction process. The reagents adopted by the extracting agent are all commonly used chemical reagents in a laboratory, and are low in cost, so that the cost is reduced; and the extracting agent is stable in component, does not need to be prepared when needed,can be repeatedly used, and can be stored at room temperature for at least 3 months. At the same time, the process of extracting the trace amount of DNA of insect samples is simple and rapid, and theextraction can be completed within 30 s. The DNA extracted by the extracting agent has stable quality and can be directly used for subsequent experimental detection. The biggest advantage of the method is that only one extracting agent is used, the extraction process can be completed within 30 s, and the extracted DNA has stable quality and can be directly applied to subsequent experimental detection.

Description

Technical field [0001] The patent of the present invention relates to the field of biotechnology, in particular to an extractant for extracting trace DNA from insects and its application. Background technique [0002] With the development of modern molecular biology and informatics, various DNA-based gene detection technologies have developed rapidly, such as allele specific amplification (ASA), restriction fragment length polymorphism analysis ( Restrietion fragment length polymorphism (RFLP), PCR single / double strand conformation polymorphism (SSCP), random amplified polymorphic DNA (RAPD), loop-mediated isothermal Amplification technology (Loop-mediated isothermal amplification, LAMP), etc., are widely used in the fields of insect species identification and identification, resistance level monitoring, insect biological characteristics research, etc., for the protection of insect biodiversity, scientific management of harmful insects, and beneficial insects in China Reasonable...

Claims

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Application Information

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IPC IPC(8): C12N15/10
CPCC12N15/1003
Inventor 蒋红云穆罕默德·欧麦·西尔赵真真张兰张燕宁毛连纲
Owner INST OF PLANT PROTECTION CHINESE ACAD OF AGRI SCI
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