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Kit and extraction method for quickly extracting microbial genome DNA from animal fecal microorganisms

A technology of animal feces and extraction methods, applied in the biological field, can solve the problems of inability to identify bacteria at the species level, a large amount of labor and time, and achieve the effects of saving extraction time and simplifying experimental steps

Inactive Publication Date: 2015-04-22
FUJIAN NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There are many ways to identify bacteria in feces, commonly used are: bacteriological isolation and culture identification technology, biochemical test identification, morphological identification and analysis, etc. These methods require a lot of labor and time, and sometimes cannot identify bacteria at the species level

Method used

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  • Kit and extraction method for quickly extracting microbial genome DNA from animal fecal microorganisms

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Effect test

Embodiment 1

[0054] A kit for rapidly extracting microbial genomic DNA from animal feces, which consists of a DNA adsorption column and the following working solution:

[0055] Working solution A: composed of potassium dihydrogen phosphate with a final concentration of 0.25mol / L and sodium hydroxide with a final concentration of 0.175mol / L, pH=7.2;

[0056] Working solution B: composed of tris (Tris) at a final concentration of 10mmol / L and ethylenediaminetetraacetic acid (EDTA) at a final concentration of 1mmol / L, pH=8.0;

[0057] Working solution C: 10mg / ml lysozyme;

[0058] Working solution D: 20mg / ml proteinase K;

[0059] Working solution E: sodium dodecyl sulfate (SDS) with a mass concentration of 10%;

[0060] Working solution F: the volume ratio is a mixed solution of phenol: chloroform: isoamyl alcohol=25:24:1;

[0061] Working solution G: the volume ratio is a mixed solution of chloroform:isoamyl alcohol=24:1;

[0062] Precipitating agent: mass concentration 70% ethanol;

...

Embodiment 2

[0068] A kit for rapidly extracting microbial genomic DNA from animal feces, which consists of a DNA adsorption column and the following working solution:

[0069] Working solution A: composed of potassium dihydrogen phosphate with a final concentration of 0.30mol / L and sodium hydroxide with a final concentration of 0.185mol / L, pH=7;

[0070] Working solution B: composed of tris (Tris) at a final concentration of 15mmol / L and ethylenediaminetetraacetic acid (EDTA) at a final concentration of 5mmol / L, pH=6.5;

[0071] Working solution C: 30mg / ml lysozyme;

[0072] Working solution D: 35mg / ml proteinase K;

[0073] Working solution E: sodium dodecyl sulfate (SDS) with a mass concentration of 10-20%;

[0074] Working solution F: the volume ratio is a mixed solution of phenol: chloroform: isoamyl alcohol=25:24:1;

[0075] Working solution G: the volume ratio is a mixed solution of chloroform:isoamyl alcohol=24:1;

[0076] Precipitating agent: mass concentration 75% ethanol;

...

Embodiment 3

[0082] A kit for rapidly extracting microbial genomic DNA from animal feces, which consists of a DNA adsorption column and the following working solution:

[0083] Working solution A: composed of potassium dihydrogen phosphate with a final concentration of 0.37mol / L and sodium hydroxide with a final concentration of 0.2mol / L, pH=8;

[0084] Working solution B: composed of tris (Tris) at a final concentration of 20mmol / L and ethylenediaminetetraacetic acid (EDTA) at a final concentration of 10mmol / L, pH=5.0;

[0085] Working solution C: 50mg / ml lysozyme;

[0086] Working solution D: 50mg / ml proteinase K;

[0087] Working solution E: sodium dodecyl sulfate (SDS) with a mass concentration of 10-20%;

[0088] Working solution F: the volume ratio is a mixed solution of phenol: chloroform: isoamyl alcohol=25:24:1;

[0089] Working solution G: the volume ratio is a mixed solution of chloroform:isoamyl alcohol=24:1;

[0090] Precipitating agent: mass concentration 80% ethanol;

...

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Abstract

The invention discloses a kit and an extraction method for quickly extracting microbial genome DNA from animal fecal microorganisms. The extraction method for extracting microbial genome DNA from animal fecal microorganisms comprises the following five steps: pre-treatment of feces; splitting decomposition of microbial cells in feces; dissociation of DNA and abstraction of impure proteins; DNA adsorption by a DNA adsorption column and removal of impurities; and elution of DNA. The kit disclosed by the invention is capable of absorbing the genome DNA by a silicone mold adsorption column and removing impurities such as proteins by using special deproteinization liquid and wash liquid. Compared with conventional extraction methods for extracting microbial genome DNA from animal fecal microorganisms, the microbial genome DNA from animal fecal microorganisms can be extracted within 4 hours, so that not only is a lot of extraction time shortened, but also the experimental steps are simplified. Therefore, the microbial genome DNA from animal fecal microorganisms can be effectively, quickly and economically extracted.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a kit and an extraction method for rapidly extracting animal feces microbial genome DNA. Background technique [0002] Animal intestines are colonized by a group of very complex and relatively stable microorganisms, which form an extremely complex micro-ecosystem and have a very important impact on animal health. The change of intestinal microecology has important significance and influence on the body. The study of intestinal microecology, especially the dominant flora, is attracting more and more attention, and has become a current research hotspot. Due to the long length of the intestinal tract, endoscopic Obtaining intestinal flora specimens in stages has not been implemented clinically due to invasiveness (trauma caused by mechanical sampling), difficulty, and time-consuming and labor-intensive factors. Analysis of the flora in feces can indirectly reflect the intest...

Claims

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Application Information

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IPC IPC(8): C12N15/10
Inventor 王清水余彦
Owner FUJIAN NORMAL UNIV
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