Method for efficiently amplifying microbial DNA from soil samples

Abstract

The invention relates to the technical field of molecular biology, and discloses a method for efficiently amplifying microbial DNA from soil samples. The method includes the following steps that step1, the soil samples are collected; step 2, microbial nucleic acid extraction is carried out on the collected soil samples; step 3, quality inspection is carried out on extracted nucleic acid; step 4,the amplification effects of an original PCR system and an optimized PCR system are compared; and step 5, qPCR verification is carried out, specifically, the concentration of the best bovine serum albumin (BSA) in an amplification system is fortified. By collecting special soil samples with high content of humic acid, organic matter, heavy metal ions and the like, and after successful extraction with a Spin Kit For Soil, PCR amplification fails. By optimizing the PCR and qPCR systems, adding bovine serum albumin (BSA) to the PCR system and finding the optimal concentration, the amplification success rate reached 100%, and the PCR amplification efficiency is greatly improved. It is verified that adding of bovine serum albumin (BSA) to the PCR system has no effect on the soil samples that can be amplified normally.

Description

technical field [0001] The invention relates to the technical field of molecular biology, in particular to a method for efficiently amplifying microbial DNA from soil samples. Background technique [0002] Soil microorganisms are an important part of the soil ecosystem. They are the driving force of material transformation in soil: such as nitrogen fixation, nitrification, denitrification, decomposition and synthesis of humus, and promotion of soil organic matter decomposition and nutrient transformation. At present, culturable microorganisms only account for 1% of the total microorganisms in the environment, and molecular biology techniques that do not depend on cultivation techniques are widely used. In recent years, with the rapid development of next-generation high-throughput sequencing technology and the substantial reduction in cost, more and more researchers have used sequencing technology to analyze the diversity of soil microbial communities. However, the basis of ...

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Problems solved by technology

However, the soil composition is complex, containing a large amount of humic acid, heavy metal ions, etc., which are difficult to completely remove during the extraction process. The residues of these substances will not only interfere with the extraction process, but also seriously inhibit the subsequent PCR amplification, resulting in the failure of library construction and sequencing.

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