Zabarovskiĭ E R, Turina O V
Mol Biol (Mosk). 1988 Nov-Dec;22(6):1451-5.
A modified procedure in two versions (micro, for 10 ml of phage lysate, and macro, for 200-500 ml) is described for preparing lambda phage DNA. The main advantage of the modified method is that it gives a possibility to isolate high-quality DNA from lambda phage lysates in 2-3 hrs. Only standard solutions (TE, NaCl, SDS, MgCl2, EDTA, RNAse A) were used throughout the whole protocol. Incubation with DNAse I and proteinase K was omitted and in microvariant concentration of the phage by PEG 6000 was excluded. Digestion by RNAse A was performed in solution with EDTA and SDS and leads to RNA degradation. The yields of DNA (0.5-2 micrograms per ml of L-broth) are similar to those obtained by other methods. DNA quality is better than in the samples of DNA prepared by other express-methods and practically the same as after CsCl centrifugation. DNA can be used for splitting by restriction enzymes, cloning and gene library construction.
本文描述了一种改良方法的两个版本（微量版，用于10毫升噬菌体裂解液；常量版，用于200 - 500毫升）来制备λ噬菌体DNA。改良方法的主要优点是能够在2 - 3小时内从λ噬菌体裂解液中分离出高质量的DNA。在整个实验方案中仅使用了标准溶液（TE、NaCl、SDS、MgCl₂、EDTA、RNA酶A）。省略了用DNA酶I和蛋白酶K的孵育步骤，并且在微量版中排除了用PEG 6000浓缩噬菌体的步骤。RNA酶A的消化是在含有EDTA和SDS的溶液中进行的，可导致RNA降解。DNA产量（每毫升L肉汤0.5 - 2微克）与其他方法获得的产量相似。DNA质量优于通过其他快速方法制备的DNA样品，实际上与氯化铯离心后的质量相同。DNA可用于限制性内切酶切割、克隆和基因文库构建。
