A procedure for large-scale plasmid isolation without using ultracentrifugation.
An expedient procedure for large-scale plasmid isolation from Escherichia coli strains without using ultracentrifugation or special setups or reagents is described. The protocol, which utilizes a modified alkaline extraction procedure as well as differential precipitations by isopropanol and lithium chloride, is simple and rapid and yet produces plasmid DNA with a yield of about 2 mg/liter culture. The isolated plasmids consisted of mostly monomeric and dimeric covalently closed circular DNA. The plasmids could be digested by various restriction endonucleases and were compatible with gene cloning, transfection-gene expression, and viral production.
Chakrabarti
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Department of Biochemistry, School of Medicine, Meharry Medical College, Nashville, Tennessee 37208.
Sitaric
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Ohi
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D000433:1-Propanol; D011232:Chemical Precipitation; D002712:Chlorides; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification; D004587:Electrophoresis, Agar Gel; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D008094:Lithium; D018021:Lithium Chloride; D010957:Plasmids / Q000302:isolation & purification*; D014162:Transfection
D002712:Chlorides; D004269:DNA, Bacterial; D000433:1-Propanol; D008094:Lithium; D018021:Lithium Chloride