噬菌体基因组DNA快速提取方法

Development of aRapid Method for Extraction of Genomic DNAs of Bacteriophages

针对传统噬菌体基因组DNA的提取方法缺乏广泛适用性,并且提取中用到酚/氯仿抽提,操作步骤繁琐,会对操作人员身体造成一定程度的损伤等不足,笔者建立了1种噬菌体基因组DNA快速提取方法。其提取的主要步骤包括宿主核酸降解、噬菌体粒子沉淀、噬菌体DNA释放、杂质去除、DNA吸附、DNA清洗和DNA洗脱等。结果表明,通过此法能快速提取以溶藻弧菌、大肠杆菌和阴沟肠杆菌为宿主菌的多种噬菌体基因组DNA,最快可以在2 h内完成,而且提取的产量和纯度均较高,DNA浓度超过100 ng·μL^(-1),OD_(260/280)达到1.8以上,提取产物不受宿主菌基因组DNA污染,可用于下游的实验分析。

Molecular biologicresearch on bacteriophages is primarily based on genomic DNA extraction. The traditional DNA extraction for bacteriophages is complicated in operation andthe use of phenol and chloroform in the DNA extraction is harmful to operator. A new rapid method for bacteriophage genome DNA extraction was developed. Procedures in the method includes nucleic acid degradation of host strains,bacteriophage precipitation,bacteriophage DNA release,impurity removal DNA adsorption,DNA washing,and DNA elution. The results indicated that bacteriophagegenomic DNAs,the host strains of which are Vibrio alginolyticus,Escherichia coli and Enterobacter cloacae,respectively,were extractedin two hours at the soonest,with the yield and purity of genomic DNAsbeing up to 100 ng·μL^-1 and 1. 8（ OD260/280）,respectively. This extraction method can prevent the DNA contamination from the host strains,and thus the extractedgenomic DNA of bacteriophagescan be used in further experiments.