A new rapid procedure for the preparation of plasmid DNA |
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| Affiliation: | 1. Microbial Technology Laboratory, PHLS Centre for Applied Microbiology & Research, Porton Down, Salisbury, Wiltshire SP4 0JG, United Kingdom;2. The Biotechnology Centre, University of Cambridge, Downing Street, Cambridge CB2 3EF, United Kingdom;3. Agricultural Division, Imperial Chemical Industries plc., Billingham, Cleveland, United Kingdom;1. Department of Internal Medicine, Division of Neurology, Showa University School of Medicine, Tokyo, Japan;2. Department of Pharmacology, Showa University School of Medicine, Tokyo, Japan;1. Department of Radiology, West China Hospital, Sichuan University, 37# Guo Xue Xiang, Chengdu, Sichuan 610041, China;2. Department of Radiology, West China Second University Hospital, Sichuan University, 20# Section 3 South Renmin Road, Chengdu, Sichuan 610041, China;1. Hubei Key Laboratory of Economic Forest Germplasm Improvement and Resources Comprehensive Utilization, Hubei Collaborative Innovation Center for Characteristic Resources Exploitation of Dabieshan, Huanggang Normal University, Hubei, China;2. Department of Food Science, Rutgers University, New Brunswick, NJ 08901, USA;3. Department of Chemical Biology, Rutgers-The State University of New Jersey, Piscataway, NJ 08854, USA |
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| Abstract: | This report describes a simple and efficient procedure for the isolation of plasmid DNA free from chromosomal DNA, cellular RNA, and protein. The technique comprises a modified cleared lysate procedure of D. B. Clewell and D. R. Helinski (1969, Proc. Natl. Acad. Sci. USA, 62, 1159–1166) followed by high-performance liquid chromatography on a Dupont Bioseries GF250 surface stable diol-coated silica gel permeation column (Zorbax) for the final purification of the plasmid DNA. The use of HPLC facilitates rapid and high-resolution separations within 3–4 h. Plasmid DNA produced in this manner retains its biological activity and exhibits yields equal to those obtained by the conventional cesium chloride-ethidium bromide density centrifugation method. |
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