Method for simultaneously extracting total DNA and RNA from compost

Abstract

The invention discloses a method for simultaneously extracting total DNA and RNA from compost, which comprises the following steps of: adding corrosion removal buffer solution into a compost sample, shaking and uniformly mixing the solution and the compost, standing the mixture, and centrifuging the suspension; repeating the operation till the centrifuged supernatant is clarified; grinding the sample after corrosion removal by using liquid nitrogen till the sample is powdery, then placing the powdery sample in cracking solution, shaking and uniformly mixing the sample and the solution, putting the mixture in an ice bath, and centrifuging the mixture; taking the cracked supernatant, adding sodium acetate and extraction buffer solution into the supernatant, and centrifuging the mixture after shaking; adding chloroform-isoamylol into the extracted upper aqueous phase, and centrifuging the mixture after shaking; adding isopropanol into the mixture to settle RNA, and washing, drying and dissolving the settled RNA to obtain purified RNA; and adding Tris alkali solution into the extracted organic phase, adding chloroform-isoamylol into the centrifuged upper aqueous phase, centrifuging the mixture, then adding sodium acetate and anhydrous ethanol into the mixture to settle DNA, and washing and dissolving the mixture to obtain purified DNA. The method has the advantages of large yield, good quality, low cost and the like.

Description

technical field [0001] The present invention relates to a method for extracting DNA and RNA, in particular to a method for extracting total DNA and RNA in compost. Background technique [0002] Composting refers to the microbiological process of artificially promoting the transformation of degradable organic matter into stable humus by using microorganisms such as bacteria, actinomycetes, fungi and other widely distributed in nature or artificially added special bacteria. It can be seen that microorganisms are the main body of the composting process, so a full understanding of the microbiological principles can provide a basis for improving the composting process and improving the efficiency of composting. The traditional methods of microbial research on compost are mainly microbial culture and purebred separation technology. Because the vast majority of microorganisms (>95%) in the environment cannot be studied by culture, and the microorganisms in compost are diverse an...

AI Technical Summary

Problems solved by technology

Due to the wide variety of microorganisms in compost and the complexity of compost components (often containing a large amount of impurities, such as humus, polysaccharides, polyphenols, heavy metals, etc.), this brings great obstacles to the extraction and purification of DNA and RNA.

Ribonucleases exist widely in the environment, which makes the extraction of RNA more difficult. Therefore, conventional DNA and RNA extraction and purification methods cannot be directly applied to composting microbial environments. How to fully lyse microbial cells and release DNA With RNA, it can effectively reduce the pollution of humus and other impurities, so as to obtain high yield and high purity DNA and RNA, which is the key to molecular ecology research on compost microbial communities

In addition, most of the separation and purification of DNA and RNA in traditional methods are carried out separately. There are few methods for extracting DNA and RNA at the same time, and the purity is also low. It is difficult to meet the requirements of downstream experiments, and the cost of using kits is very high. high

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