一种制备适用于测序的高质量λ噬菌体DNA的快速新方法。

Manfioletti G, Schneider C

European Molecular Biology Laboratory, Heidelberg, FRG.

Nucleic Acids Res. 1988 Apr 11;16(7):2873-84. doi: 10.1093/nar/16.7.2873.

A method is described for the rapid purification of high quality lambda DNA. The method can be used from either liquid or plate lysates and on a small scale or a large scale. It relies on the preadsobtion of all polyanions present in the lysate to an "insoluble" anion-exchange matrix (DEAE or TEAE). Phage particles are then disrupted by combined treatment with EDTA/proteinase K and the resulting DNA is precipitated by the addition of the cationic detergent cetyl (or hexadecyl)-trimethyl ammonium bromide-CTAB ("soluble" anion-exchange matrix). The precipitated CTAB-DNA complex is then exchanged to Na-DNA and ethanol precipitated. The resultant purified DNA is suitable for enzymatic reactions and provides a high quality template for dideoxy-sequence analysis.

描述了一种快速纯化高质量λ DNA的方法。该方法可用于液体或平板裂解物，可小规模或大规模使用。它依赖于裂解物中存在的所有聚阴离子预先吸附到“不溶性”阴离子交换基质（DEAE或TEAE）上。然后通过EDTA/蛋白酶K联合处理破坏噬菌体颗粒，并通过加入阳离子去污剂十六烷基（或十六烷基）三甲基溴化铵-CTAB（“可溶性”阴离子交换基质）沉淀得到的DNA。然后将沉淀的CTAB-DNA复合物交换为Na-DNA并乙醇沉淀。所得纯化的DNA适用于酶促反应，并为双脱氧序列分析提供高质量模板。