Microbial nucleic acid extraction method with host genome DNA removal function and kit

Abstract

The invention discloses a microbial nucleic acid extraction method with a host genome DNA removal function and a kit. The invention provides a complete set of primer pairs, including one or more primer pairs. Each primer pair correspondingly amplifies one highly repetitive sequence in host genome DNA, wherein the highly repetitive sequence is selected from at least one gene in the following gene family: the gene family includes an MIR family gene and / or an ALu family gene. The microbial DNA extraction method with the host genome DNA removal function has the advantage that the microbial genomeDNA enrichment effect is stronger. According to the method, the host genome DNA is removed after the nucleic acid is released through lysis, then the microbial genome DNA is enriched for purificationand recovery, and meanwhile, viral nucleic acid in host cells can be enriched without changing the metagenome flora structure.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a microbial nucleic acid extraction method and a kit capable of removing host genome DNA. Background technique [0002] Metagenomics is also called microbial environmental genomics and metagenomics. It mainly constructs genomic libraries by directly extracting the genomic DNA of all microorganisms from environmental samples, and uses genomics research strategies to study all microorganisms contained in environmental samples. With the rapid development of next-generation sequencing technology, metagenomics research has been applied to various fields, especially in the field of clinical medicine in recent years, including infectious disease diagnosis, pathogen drug sensitivity, infectious disease Epidemiology, epidemiology, etc., have vigorously promoted the development of precision medicine for infectious diseases and improved the level of clinical diagnosis and treatment ...

AI Technical Summary

Problems solved by technology

[0005] However, Qiagen's QIAamp DNA Microbiome Kit and Meiji Bio's blood-enriched bacterial DNA extraction kit (D3148-02) are both based on the principle that chemical reagents preferentially crack and release host genomic DNA to achieve the purpose of removing host genomic DNA, but due to Due to the diversity of bacterial species, it is difficult to ensure that the release of host genomic DNA by chemical reagents will not affect the microbial community structure in the sample. At the same time, most of the microorganisms such as DNA viruses and RNA viruses in the sample are integrated into the host genome or free in the host. It exists in the form of cytoplasm, and removes the viral genomic DNA and RNA while removing the host genomic DNA

This method cannot extract viral DNA and RNA in the sample, so that such products often miss viral pathogens in metagenomic research, and the practicability is greatly reduced. Therefore, it is difficult to remove the host genome DNA only by chemical and physical methods. The purpose of preserving all metagenomic DNA at the same time

[0006] The patent "Metagenome Extraction Method" (authorized number: CN103060309B) provides a brand-new way to remove host genomic DNA, but the probe sequence provided in the authorized patent is not derived from the repetitive sequence Alu sequence, and the patent uses gene synthesis The method of preparing capture probes is expensive and not suitable for direct application in nucleic acid extraction kits

The capture hybridization probe is only designed for the repetitive sequence Alu sequence, and the probe is prepared by gene synthesis. Due to the limitation of the synthesis cost, the length of the synthesized probe generally does not exceed 50bp, and the binding efficiency is low due to the length limitation. Therefore, the patent provides The method has technical shortcomings such as fewer sites, low binding efficiency, and high cost.

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