Method for extracting total DNAs of soil microorganisms at high purity

Abstract

The invention discloses a method for extracting total DNAs of soil microorganisms at high purity. CTAB (Cetyltrimethyl Ammonium Bromide), lysozyme, protease and SDS (Sodium Dodecyl Sulfonate) act together to perform lysis of cells and simultaneously freeze and thaw the cells repeatedly; therefore, an excellent cell lysis effect is achieved; the humic acid is removed by utilizing PVP (Polyvinyl Pyrrolidone), CaC12 and BSA (Bull Serum Albumin); and the DNAs are precipitated by utilizing isopropanol so that the purity of the obtained DNAs is high. The method is simple and convenient; and without being purified, the DNAs obtained by the method can be used for subsequent PCR (Polymerase Chain Reaction) analysis and DGGE (Denaturing Gradient Gel Electrophoresis) analysis. The purity of the extracted DNAs meets the requirement for directly performing subsequent molecular biological research; the extracted DNA segments can be subjected to 16SrDNA amplification and DGGE atlas analysis; glue cutting recovery and basic group sequencing can be performed on specific stripes of the DGGE band; unknown bacteria can be identified through comparison with the conventional sequences in an RDP (Ribosomal Database Project) database or by establishing a new sequence probe; and therefore, the structural composition of the microbial community can be determined, and a foundation can be laid for researching the structural diversity and ecological functions of the soil microorganisms in the rhizosphere of the mulberry in future.

Description

technical field

[0001] The invention relates to the technical field of molecular ecology, in particular to a method for extracting total DNA of soil microorganisms in mulberry gardens. Background technique

[0002] For a long time, the research on the diversity of soil microorganisms has been achieved through isolation and culture. However, the vast majority of microbial populations cannot be purely cultured, which has become a limiting factor in the study of soil microbial diversity. With the continuous deepening of the research on the structure of soil microbial communities, modern molecular biology research methods such as PCR-RFLP, PCR-SSCP, PCR-DGGE, etc. have avoided the shortcomings of traditional research methods that cannot fully obtain soil microbial diversity information, and can The diversity of soil microbial DNA truly reflects the structure of the soil microbial population, and the efficient and high-quality extraction of the total DNA of the soil microbial co...

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