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Reagent for extracting total RNA from animal tissues and extracting method

A technology of animal tissue and extraction method, applied in the direction of recombinant DNA technology, DNA preparation, etc., can solve the problems of high cost of ordinary molecular experiments, expensive RNA extraction kits, etc., and achieve high purity of total RNA products, low cost, and simple operation process Effect

Inactive Publication Date: 2010-09-15
SHANXI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The RNA extraction kits (such as TRIzol) on the market are extremely expensive, bringing huge costs to common molecular experiments

Method used

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  • Reagent for extracting total RNA from animal tissues and extracting method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0011] Mix 40 g of guanidine acetic acid, 0.1 g of sodium lauryl sulfate, 5 g of phenol and 100 g of sterilized distilled water, and adjust the pH to 5.5 with glacial acetic acid.

Embodiment 2

[0013] Mix 350 g of guanidine acetic acid, 2 g of sodium lauryl sulfate, 60 g of phenol and 1000 g of sterilized distilled water, and adjust the pH to 5 with glacial acetic acid.

[0014] The following is an example of using the extraction reagent of the present invention to extract total RNA:

Embodiment 3

[0016] Add 450μl of the reagent of Example 1 into a centrifuge tube sterilized by high temperature, add 50mg of crab gill tissue to the centrifuge tube, homogenize in an ice bath until no particles are transparent, and let stand at room temperature for 10 minutes; centrifuge at 10000 rpm for 2 minutes and aspirate the supernatant , Transfer the supernatant to another centrifuge tube; add 1 / 3 volume of the supernatant chloroform, shake and mix; centrifuge at 10000rpm for 3min, aspirate the supernatant, and transfer the supernatant to the third centrifuge tube; add the supernatant 1 / 2 volume of 20% Li 2 SO 4 Mix well with 1 / 2 volume of frozen isopropanol, and let stand at -80°C for 10 minutes. Centrifuge at 10000rpm for 2min, recover the precipitate, wash the precipitate with 75% ethanol to obtain total RNA.

[0017] The above total RNA precipitate was dissolved in 100 μl of RNase-free water.

[0018] Electrophoresis detection: the electrophoresis buffer is 0.5×TBE buffer 1L, add 3g...

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Abstract

The invention provides a reagent for extracting total RNA from animal tissues, which comprises the following components in part by weight: 30 to 40 parts of glucocyamine, 0.1 to 0.3 part of lauryl sodium sulfate, 5 to 10 parts of phenol and 100 parts of sterilized distilled water. The pH of the reagent is adjusted to between 5 and 6 by using glacial acetic acid. The invention provides a method for extracting the total RNA from the animal tissues, which uses Li ions and isopropanol to deposit the total RNA. The operation is simple in the whole process, the cost is low, and the obtained RNA has high purity and good stability.

Description

Technical field [0001] The invention relates to molecular biotechnology, in particular to a reagent and an extraction method for extracting total RNA from animal tissues. Background technique [0002] RNA extraction technology is not only an important part of molecular biology technology, but also an important foundation of functional genomics research technology. Studying the regulation mechanism of genes in organisms from the RNA level has become an important means of molecular biology research. Many molecular biotechnologies, including RNA mapping, Northern blotting, nuclease protection test, PT-PCR (reverse transcription polymerase chain reaction), real time PCR, cDNA library construction, and other experiments, largely depend on the quality of RNA. Because RNA is unstable, it is more difficult to extract than DNA. The key to the success of the experiment is whether there is RNase contamination. When extracting RNA, avoid the pollution of exogenous RNase and inhibit the ac...

Claims

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Application Information

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IPC IPC(8): C12N15/10
Inventor 高爱保王兰袁慧
Owner SHANXI UNIV