Summary
A rapid and easy final purification method is described for the isolation of plasmids from B. fragilis. Using RbTCA density gradient centrifugation in an airfuge ultracentrifuge ccc plasmid DNA can be separated from RNA, residual chromosomal DNA, linear and oc plasmid DNA. Pure ccc plasmid DNA is obtained from cultures of between 1 ml and 2 l in less than one day.
References
Birnboim HC, Doly J (1979) A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res 7:1513–1523
Burke RL, Bauer WR (1977) The properies of native and denatured DNA in buoyant rubidium trichloroacetate at neutral pH. Nucleic Acids Res 4:1891–1909
Burke RL, Bauer WR (1978) The helix-coil transition of closed and nicked DNAs in aqueous neutral trichloroacetate solutions. Nucleic Acids Res 5:4819–4836
Burke RL, Bauer WR (1980) The early melting of closed duplex DNA: analysis by banding in buoyant neutral rubidium trichloroacetate. Nucleic Acids Res 8:1145–1165
Clewell DB, Helinski DR (1969) Supercoiled circular DNA-protein complex in Escherichia coli: Purification and induced conversion to an open circular DNA form. Biochemistry 62:1157–1166
Colman A, Byers MJ, Primose SB, Lyons A (1978) Rapid purification of plasmid DNAs by hydroxyapatite chromatography. Eur J Biochem 91:303–310
Cornelis P, Digneffe C, Willemot K, Colson C (1981) Purification of Escherichia coli amplificable plasmids by highsalt sepharose chromatography. Plasmid 5:221–223
Guerry P, LeBlanc DJ, Falkow S (1973) General method for the isolation of plasmid deoxyribonucleic acid. J. Bacteriol 116:1064–1066
Guiney DG, Davies CE (1975) Isolation of plasmid deoxyribonucleic acid from two strains of bacteroides. J Bacteriol 124:503–510
Guiney DG, Davies CE (1978) Identification of a conjugative R plasmid in Bacteroides ochraceus capable of transfer to Escherichia coli. Nature 274:181–182
Hansen JB, Olsen RH (1978) Isolation of large bacterial plasmids and characterization of the P2 incompatibility group plasmids pMGl and pMG5. J Bacteriol 135:227–238
Holmes DS, Quigley M (1981) A rapid boiling method for the preparation of bacterial plasmids. Anal Biochem 114:193–197
Kado CI, Liu ST (1981) Rapid procedure for detection and isolation of large and small plasmids. J Bacteriol 145:1365–1373
Klein RD, Selsing E, Wells RD (1980) A rapid microscale technique for isolation of recombinant plasmid DNA suitable for restriction enzyme analysis. Plasmid 3:88–91
Mays TD, Smith CJ, Welch RA, Delfini C, Macrina L (1982) Novel antibiotic resistance transfer in Bacteroides. Antimicrob Agents Chemother 21:110–118
Meyers JA, Sanchez D, Elwell LP, Falkow S (1976) Simple agarose gel electrophoretic method for the identification and characterization of plasmid deoxyribonucleic acid. J Bacteriol 127:1529–1537
Pakroppa W, Goebel W, Müller W (1975) Isolation of closedcircular duplex DNA by chromatography on hydroxyapatite in the presence of ethidium bromide. Anal Biochem 67:372–383
Palchaudhuri S, Bawdon RE (1980) Plasmid mediated clindamycin resistance in Bacterioides fragilis. In: Stuttgard C, Rozee KR (eds) Plasmids and transposons. Academic Press, New York, London, pp 129–136
Privitera G, Botta G, Sebald M (1981) Macrolide, lincosamide, streptogramin and tetracycline transferable resistance in Bacteroides fragilis group. J Antimicrob Chemother 8:Suppl D 87–94
Ranhand FM (1982) A simple method for the qualitative demonstration of large and small plasmid deoxyribonuceic acids in gram-positive and gram-negative bacteria. Anal Biochem 122:426–432
Rashtchian A, Dubes GR, Booth SJ (1982) Tetracycline-inducible transfer of tetracycline resistance in Bacteroides fragilis in the absence of detectable plasmid DNA. J Bacteriol 150:141–147
Rashtchian A, Booth SJ (1981) Stability in Escherichia coli of an antibiotic resistance plasmid from Bacteroides fragilis. J Bacteriol 146:121–127
Rotimi VO, Duerden BI, Hafiz S (1981) Transferable plasmidmediated antibiotic resistance in Bacteroides. J Med Microbiol 14:359–370
Sharp PA, Hsu M-T, Ohtsubo E, Davidson N (1972) Electron microscope heteroduplex studies of sequence relations among plasmids of Escherichia coli. J Mol Biol 71:471–497
Stiffler PW, Keller R, Traub T (1974) Isolation and characterisation of several cryptic plasmids from clinical isolates of Bacteroids fragilis. J Infect Dis 130:544–548
Stougaard P, Molin S (1981) Vertical dye-buoyant density gradients for rapid analysis and preparation of plasmid DNA. Anal Biochem 118:191–193
Talley FP, Snydman DR, Gorbach S, Malamy M (1979) Plasmid mediated, transferable resistance to clindamycin and erythromycin in Bacteroides fragilis. J Infect Dis 139:83–88
Tinnel WH, Macrina FL (1976) Extrachromosomal elements in a variety of strains representing the Bacteroides fragilis group of organisms. Infect Immun 14:955–964
Wallace BL, Bradley JE, Rogolsky M (1981) Plasmid analyses in clinical isolates of Bacteroides fragilis and other Bacterioides species. J Clin Microbiol 14:383–388
Welch R, Jones K, Macrina FL (1979) Transferable lincosamidemacrolide resistance in Bacteroides. Plasmid 2:261–268
Welch RA, Macrina FL (1981) Physical characterisation of Bacteroides fragilis R plasmid pBF4, J Bacteriol 145:867–872
Welch RA, Macrina FL (1980) Deletion analysis of a transferable Bacteroides R plasmid. In: Shuttard C, Rozee KR (eds) Plasmids and transposons. Academic Press, New-York, London, pp 137–144
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Communicated by E. Bautz
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Zöllner, E.J., Eichel-Streiber, C.v. & Schwindling, FP. Isolation and purification of plasmids from Bacteriodes fragilis using rubidium trichloroacetate density gradient centrifugation. Molec Gen Genet 189, 169–171 (1983). https://doi.org/10.1007/BF00326072
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DOI: https://doi.org/10.1007/BF00326072