Rapid and simple method for purification of nucleic acids

R Boom, CJ Sol, MM Salimans, CL Jansen… - Journal of clinical …, 1990 - Am Soc Microbiol
R Boom, CJ Sol, MM Salimans, CL Jansen, PM Wertheim-van Dillen, J Van der Noordaa
Journal of clinical microbiology, 1990Am Soc Microbiol
We have developed a simple, rapid, and reliable protocol for the small-scale purification of
DNA and RNA from, eg, human serum and urine. The method is based on the lysing and
nuclease-inactivating properties of the chaotropic agent guanidinium thiocyanate together
with the nucleic acid-binding properties of silica particles or diatoms in the presence of this
agent. By using size-fractionated silica particles, nucleic acids (covalently closed circular,
relaxed circular, and linear double-stranded DNA; single-stranded DNA; and rRNA) could …
We have developed a simple, rapid, and reliable protocol for the small-scale purification of DNA and RNA from, e.g., human serum and urine. The method is based on the lysing and nuclease-inactivating properties of the chaotropic agent guanidinium thiocyanate together with the nucleic acid-binding properties of silica particles or diatoms in the presence of this agent. By using size-fractionated silica particles, nucleic acids (covalently closed circular, relaxed circular, and linear double-stranded DNA; single-stranded DNA; and rRNA) could be purified from 12 different specimens in less than 1 h and were recovered in the initial reaction vessel. Purified DNA (although significantly sheared) was a good substrate for restriction endonucleases and DNA ligase and was recovered with high yields (usually over 50%) from the picogram to the microgram level. Copurified rRNA was recovered almost undegraded. Substituting size-fractionated silica particles for diatoms (the fossilized cell walls of unicellular algae) allowed for the purification of microgram amounts of genomic DNA, plasmid DNA, and rRNA from cell-rich sources, as exemplified for pathogenic gram-negative bacteria. In this paper, we show representative experiments illustrating some characteristics of the procedure which may have wide application in clinical microbiology.
American Society for Microbiology