冻融法快速提取真菌微量培养物基因组DNA

Rapid Isolation of Genomic DNA from Tiny Culture Based on Freezing and Thawing

目的:外生菌根真菌松茸Tricholoma matsutake(S.Ito et Imai)Sing.又名松口蘑,是一种极其珍稀昂贵的野生食用蘑菇资源。为了保护利用松茸资源的生物多样性,需要研究发展适用松茸的基因组DNA的提取方法。方法:对传统的液氮冷冻研磨的细胞破壁方法加以改进,采用液氮冷冻与室温融化交替处理松茸菌丝体与其它供试真菌细胞,进而萃取、浓缩、沉淀DNA。结果:利用本文研究的方法分离纯化出了满足分子生物学实验要求的高质量细胞总DNA。结论:该法相对简便、安全,可行、可靠,对供试真菌培养物需求量较少,既适用于担子真菌松茸,也适用于供试的其它真菌类群如酵母菌、丝状霉菌,应用前景广阔。

Objective：Tricholoma matsutake（S.Ito et Imai） Sing.is an ectomycorrhizal fungus producing economically important mushroom.The successful extraction of DNA from fungal isolates is essential to the investigation of genetic diversity of T.matsutake resources on the brink of annihilation.Method：A innovatory method for rapid isolation of genomic DNA from tiny mycelia or cell culture was developed in this paper,with fungal cell walls broken based on freezing and thawing technology.Result：High molecular quality of DNAs was prepared with good integrality and satisfied the meets of routine molecular biology experiments.Conclusion： The method developed is effective and fit for fungi tested included basidiomycete,yeast,and filamentous fungus.