Chang P K, Natori M
Department of Applied Biological Science, College of Agriculture and Veterinary Medicine, Nihon University, Fujisawa, Japan.
J Biotechnol. 1994 Aug 31;36(3):247-51. doi: 10.1016/0168-1656(94)90156-2.
We have established two simple, reliable, economical and time-saving methods, one for plasmid and phagemid DNAs another for bacteriophage DNAs, that can be applied for any desired scale of vector and its recombinant DNAs preparation. The methods require neither toxic chemicals nor expensive enzymes or chemical reagents. In case of the small-scale preparation, the entire procedure can be done in one microfuge tube. The A260/A280 values for isolated DNAs were constantly between 1.60 and 1.85. The isolated plasmid or phagemid DNA can be used for restriction digestion, religation, transformation, construction of deletion mutants, sequencing, PCR and in vitro transcription. The double-stranded and single-stranded phage DNAs prepared from the present method have the quality to serve as good templates for PCR and site-directed mutagenesis experiments.
我们建立了两种简单、可靠、经济且省时的方法，一种用于制备质粒和噬菌粒DNA，另一种用于制备噬菌体DNA，可应用于任何所需规模的载体及其重组DNA的制备。这些方法既不需要有毒化学物质，也不需要昂贵的酶或化学试剂。对于小规模制备，整个过程可以在一个微量离心管中完成。分离出的DNA的A260/A280值始终在1.60至1.85之间。分离出的质粒或噬菌粒DNA可用于限制性消化、重新连接、转化、缺失突变体构建、测序、PCR和体外转录。用本方法制备的双链和单链噬菌体DNA具有作为PCR和定点诱变实验良好模板的质量。
