Ishaq M, Wolf B, Ritter C
Dept. of Pathobiology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia 19104-6049.
Biotechniques. 1990 Jul;9(1):19-20, 22, 24.
A rapid procedure for the large-scale isolation of plasmid DNA is described. The method utilizes cetyltrimethylammonium bromide to precipitate the plasmid following extraction of DNA by lysozyme digestion and boiling. The plasmid is then purified by passing through the spin column pZ523. The purity and yield of the plasmid obtained with this method is similar to that isolated by cesium chloride-ethidium bromide gradient centrifugation. The method does not involve any phenol-chloroform extractions and takes five to six hours for completion after growth of the bacterial cells. The plasmid obtained is amenable to digestion with various restriction endonucleases, can be used for cloning with high efficiency and is also suitable as template for dideoxy sequencing.
本文描述了一种大规模分离质粒DNA的快速方法。该方法利用十六烷基三甲基溴化铵在通过溶菌酶消化和煮沸提取DNA后沉淀质粒。然后通过pZ523旋转柱纯化质粒。用该方法获得的质粒的纯度和产量与通过氯化铯-溴化乙锭梯度离心分离的质粒相似。该方法不涉及任何酚-氯仿萃取，在细菌细胞生长后完成整个过程需要五到六个小时。所获得的质粒适合用各种限制性内切酶进行消化，可高效用于克隆，也适合作为双脱氧测序的模板。
