一种改良的可直接用于PCR扩增的土壤DNA提取方法

An Improved Soil DNA Extraction Method for Direct PCR Amplification

本研究对Zhou氏土壤DNA抽提方法进行了改良,减少了土壤用量、孵育和裂解时间,增加了土壤淋洗和硫酸铝处理步骤。接着比较了改良法和Zhou氏法抽提四种类型土壤DNA的效果。结果表明,将土壤用量减少至0.4 g、孵育和裂解时间分别降至20 min和30 min,极大地简易了操作过程,大幅节省了实验所需时间(约为Zhou氏法所需时间的一半)。裂解前增加淋洗步骤和裂解过程中加入硫酸铝都提高了DNA的纯度。前者同时提高了DNA得率,但后者稍微降低了DNA得率。与Zhou氏法相比较,改良法提取4种类型土壤(黑土,黄土,潮土,红土)的DNA在得率和纯度方面均得到提高,并且可以直接用于16S rRNA基因的PCR扩增等分子操作。总之,本改良法简便了DNA提取过程、大大节省了实验时间,可用于从不同类型土壤中抽提适合于PCR扩增等分子操作的DNA。

In this study,the Zhou’s DNA extraction method from soil was improved.Soil usage amount,lysozyme incubation and SDS lysis times were reduced and the steps of soil pre-washing and aluminum sulfate treatment were added in the improved method.Then,the effects of the DNA extracted from four types of soils by the modified method were compared with that of the Zhou’s method.The results showed that reducing the amount of soil to 0.4 g,the incubation and lysis time to 20 min and 30 min,respectively,greatly simplified the operation process and greatly saved the experimental time(about half the time required for Zhou’s method).Both the soil pre-washing step prior to cell lysis and aluminum sulphate treat in cell lysis increased the purity of the DNA.The former increased DNA yield,but the latter slightly reduced it.Compared with the Zhou’s method,DNA extracted by the modified method from four types of soils(black soil,loess soil,fluvo-aquic soil and red soil)were improved in yield and purity,and could be directly used for molecular biology techniques,e.g.PCR amplification of 16 S rRNA gene.In summary,this modified method could greatly simplify the DNA extraction process,drastically save the experimental time and could also be used to extract DNA suitable for molecular operations such as PCR amplification from different types of soil.