Singh Samarendra K, Banerjee Pataki C
Indian Institute of Chemical Biology, Kolkata 700032, India.
Anal Biochem. 2006 Sep 15;356(2):229-34. doi: 10.1016/j.ab.2006.06.016. Epub 2006 Jun 28.
Plasmid yield from Acidiphilium strains always had been poor following various standard methods. We adopted some simple modifications in the alkaline lysis procedure to get a better yield of plasmid from these bacteria. An approximately 10- to 20-fold increase in the plasmid yield was achieved when harvested Acidiphilium cells were preincubated 16-20 h at pH 6 in nitrogen-free medium. Another independent approach showed that freezing (-18 to -20 degrees C) of the harvested cells initially and at two subsequent steps in the alkaline lysis procedure of plasmid DNA extraction improved the yield further by 1.5- to 3-fold. The combination of these changes yielded at least 15- to 30-fold more plasmid from various Acidiphilium strains as compared with standard methods.
按照各种标准方法，嗜酸菌菌株的质粒产量一直很低。我们对碱裂解法进行了一些简单的改进，以便从这些细菌中获得更高产量的质粒。当收获的嗜酸菌细胞在无氮培养基中于pH 6下预孵育16 - 20小时时，质粒产量提高了约10至20倍。另一种独立的方法表明，在质粒DNA提取的碱裂解过程中，最初以及随后的两个步骤将收获的细胞冷冻（-18至-20摄氏度），可使产量进一步提高1.5至3倍。与标准方法相比，这些改变相结合，从各种嗜酸菌菌株中获得的质粒产量至少提高了15至30倍。
