Method for extracting plant pathogenic fungus DNA for PCR detection

Abstract

The invention provides a method for extracting plant pathogenic fungus DNA for PCR detection. Hyphal lysate is adopted for conducting heat treatment on hyphae for 10 minutes, and a template can be subjected to PCR amplification by diluted crude lysate, wherein the lysate contains 0.4-0.6wt.% SDS, 0.08-0.12M Tris-HCl with the pH value of 9.0-9.5, 0.8-1.2M NaCl and 8-12mM EDTA. The method has the advantages that the process is simpler and quicker, the cumbersome steps of liquid nitrogen grinding, centrifugation, supernatant removal and the like are omitted, the template of PCR amplification canbe obtained by directly conducting pyrolysis treatment on hyphal samples, the whole operation process is faster, and the time for extracting each sample is shorter than 15 minutes.

Description

technical field [0001] The invention belongs to the field of fungal molecular detection, and in particular relates to a method for extracting DNA of plant pathogenic fungi for PCR detection. Background technique [0002] Plant diseases caused by fungi are the most serious and widely distributed diseases in agricultural production. Molecular identification of pathogenic bacteria is an important means to quickly identify diseases and guide control work; It is an important method to understand the mechanism of disease and provide prevention and control strategies. The most commonly used technology for molecular identification and genetic transformant identification of fungi is PCR detection, the premise of which is to prepare genomic DNA as a template for PCR amplification. The traditional fungal genomic DNA preparation method requires multiple steps such as cell disruption, CTAB treatment, protein removal, DNA precipitation, and washing. At the same time, although the tradit...

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Problems solved by technology

However, there are still the following shortcomings: 1. The whole extraction process is relatively long, and it takes at least 30 minutes for a single sample; 2. It is difficult to perform liquid nitrogen grinding in a 1.5ml centrifuge tube, and it is difficult to achieve high-throughput; 3. Chelex-100 is a commercial cationic chelating resin, which needs to be purchased and is expensive, about 8000 yuan for 500g (Bio-Rad)

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