Rapid extraction kit for animal tissue genome RNA and extraction method and application

A technology of animal tissues and kits, applied in the biological field, can solve the problems of affecting amplification and sequencing, interfering with the amplification process, application limitations, etc., and achieve the effects of simplified reagent composition, broad market prospects, and good economic benefits

Inactive Publication Date: 2015-09-02
INST OF ANIMAL HUSBANDRY & VETERINARY MEDICINE HENAN ACAD OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Animal tissue contains a large amount of protein, salt and impurities, whose properties are very similar to those of nucleic acid, and it is difficult to separate them from the extracted RNA. In the process of animal tissue RNA extraction, it is easy to co-precipitate with RNA and difficult to separate , often resulting in insufficient purity and concentration of extracted RNA, which affects subsequent amplification and sequencing
[0005] In order to reduce or remove proteins, salts and impurities in anima

Method used

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  • Rapid extraction kit for animal tissue genome RNA and extraction method and application

Examples

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Effect test

Embodiment 1

[0034] Example 1: A kit and extraction method for rapid extraction of genomic RNA from animal tissues

[0035] (1) Kit reagents: including lysis solution, washing solution 1, washing solution 2, absolute ethanol and 1% (by weight) diethyl pyrocarbonate aqueous solution.

[0036] Components of each reagent:

[0037] Lysis buffer: Tris-HCl at a concentration of 50 mM, 1% (by weight) of cetyltrimethylamine bromide, 1% (by weight) of lauryl sarcosine, 5% (by weight) of Polyvinylpyrrolidone, 0.7 M sodium chloride;

[0038] Wash solution 1: EDTA with a concentration of 10 mM, 70% (volume ratio) ethanol, pH 7.0;

[0039] Wash solution 2: Tris-HCl with a concentration of 50 mM, 75% (volume ratio) ethanol, pH 7.0.

[0040] Use this kit to extract animal tissues such as heart, liver, lung, spleen, kidney, etc. These animal tissues need to be ground into a paste first, and then extracted. The extraction steps are as follows:

[0041] (1) Take 20 mg of ground animal tissue, add 500 μl...

Embodiment 2

[0048] Embodiment 2: A kit and extraction method for rapid extraction of genomic RNA from animal tissue, which is basically the same as Example 1, except that:

[0049] (1) Kit reagents: including lysis solution, washing solution 1, washing solution 2, absolute ethanol and 0.5% (by weight) diethyl pyrocarbonate aqueous solution.

[0050] Components of each reagent:

[0051] Lysis solution: 40mM Tris-HCl, 0.5% (weight ratio) cetyltrimethylamine bromide, 0.5% (weight ratio) lauryl sarcosine, 3% (weight ratio) polyvinylpyrrolidone , 0.35 M NaCl;

[0052] Wash solution 1: 5 mM EDTA, 70% (volume ratio) ethanol, pH 7.0;

[0053] Wash solution 2: 40 mM Tris-HCl, 75% (volume ratio) ethanol, pH 7.0.

[0054] The extraction steps are the same as Example 1.

Embodiment 3

[0055] Example 3: A rapid extraction kit and extraction method for genomic RNA in animal tissues

[0056] (1) The kit includes lysis solution, washing solution 1, washing solution 2, absolute ethanol and 1% (by weight) diethyl pyrocarbonate aqueous solution.

[0057] Components of each reagent:

[0058] Lysis buffer: 50 mM Tris-HCl, 1% (by weight) cetyltrimethylamine bromide, 1% (by weight) lauryl sarcosine, 5% (by weight) polyvinyl Pyrrolidone, 0.7 M NaCl;

[0059] Wash solution 1: 10 mM EDTA, 70% (volume ratio) ethanol, pH 7.0;

[0060] Washing solution 2: 50 mM Tris-HCl, 75% (volume ratio) ethanol, pH 7.0.

[0061] Follow the procedure below to extract genomic RNA from animal tissues (heart, liver, lung, spleen, kidney), whole blood, plasma, serum, animal secretions (emulsion, nasal fluid):

[0062] (1) Take 50 μl of the liquid, add 500 μl of lysate, bathe in 70°C water for 2 min, 12000 rpm for 30 s, and separate the supernatant;

[0063] (2) Transfer the supernatant t...

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Abstract

The invention relates to a rapid extraction kit for animal tissue genome RNA and an extraction method and application. The rapid extraction kit comprises lysate, flushing liquid 1, flushing liquid 2, absolute ethyl alcohol and a diethyl pyrocarbonate solution; when in extracting, solid animal tissue is milled, and liquid animal tissue is directly extracted; after a sample is cracked, the genome RNA can be absorbed to an RNA absorbing column, and then the diethyl pyrocarbonate solution is used for flushing to obtain the complete genome RNA. The kit is applicable to extraction and purifying of the genome RNA of the animal tissue; the whole extraction costs 5 minutes; the extraction efficiency is high, and the detection sensitivity is high; reliable experience basis is provided to early diagnose various RAN bacteria or viral pathogen infection. The kit is applicable to all research and development institutions, clinical pathogen detection, molecular genetics research, clinical gene diagnosis, animal disease diagnosis and disease control centers, and has a wide market prospect and high economic benefit.

Description

technical field [0001] The invention relates to a kit for rapid extraction of genome RNA from animal tissues, an extraction method and application thereof, and belongs to the field of biotechnology. Background technique [0002] The extraction and purification of RNA in molecular biology research is the first step in molecular experiments, and it is necessary to extract and purify RNA in various clinical tests or scientific research. At present, a variety of kits for detecting RNA based on RT-PCR technology have been used in clinical diagnosis and detection at home and abroad. The RNA extraction method provided by these kits is mainly the phenol-chloroform extraction method. Phenol and chloroform are denaturants in the RNA extraction process, and repeated extractions can remove proteins, salts, and impurities. However, phenol and chloroform are highly toxic to the human body and highly volatile, and the entire extraction process is cumbersome and time-consuming, which often...

Claims

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Application Information

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IPC IPC(8): C12N15/10
Inventor 李海利王克领朱文豪杨宁徐照学许峰候自花师丽刚
Owner INST OF ANIMAL HUSBANDRY & VETERINARY MEDICINE HENAN ACAD OF AGRI SCI
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