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J Bacteriol. 1996 February; 178(3): 591–599.
PMCID: PMC177699

Characterization of uncultivated prokaryotes: isolation and analysis of a 40-kilobase-pair genome fragment from a planktonic marine archaeon.

Abstract

One potential approach for characterizing uncultivated prokaryotes from natural assemblages involves genomic analysis of DNA fragments retrieved directly from naturally occurring microbial biomass. In this study, we sought to isolate large genomic fragments from a widely distributed and relatively abundant but as yet uncultivated group of prokaryotes, the planktonic marine Archaea. A fosmid DNA library was prepared from a marine picoplankton assemblage collected at a depth of 200 m in the eastern North Pacific. We identified a 38.5-kbp recombinant fosmid clone which contained an archaeal small subunit ribosomal DNA gene. Phylogenetic analyses of the small subunit rRNA sequence demonstrated it close relationship to that of previously described planktonic archaea, which form a coherent group rooted deeply within the Crenarchaeota branch of the domain Archaea. Random shotgun sequencing of subcloned fragments of the archaeal fosmid clone revealed several genes which bore highest similarity to archaeal homologs, including large subunit ribosomal DNA and translation elongation factor 2 (EF2). Analyses of the inferred amino acid sequence of archaeoplankton EF2 supported its affiliation with the Crenarchaeote subdivision of Archaea. Two gene fragments encoding proteins not previously found in Archaea were also identified: RNA helicase, responsible for the ATP-dependent alteration of RNA secondary structure, and glutamate semialdehyde aminotransferase, an enzyme involved in initial steps of heme biosynthesis. In total, our results indicate that genomic analysis of large DNA fragments retrieved from mixed microbial assemblages can provide useful perspective on the physiological potential of abundant but as yet uncultivated prokaryotes.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. [PubMed]
  • Barns SM, Fundyga RE, Jeffries MW, Pace NR. Remarkable archaeal diversity detected in a Yellowstone National Park hot spring environment. Proc Natl Acad Sci U S A. 1994 Mar 1;91(5):1609–1613. [PubMed]
  • Britschgi TB, Giovannoni SJ. Phylogenetic analysis of a natural marine bacterioplankton population by rRNA gene cloning and sequencing. Appl Environ Microbiol. 1991 Jun;57(6):1707–1713. [PMC free article] [PubMed]
  • Ceccarelli E, Bocchetta M, Creti R, Sanangelantoni AM, Tiboni O, Cammarano P. Chromosomal organization and nucleotide sequence of the genes for elongation factors EF-1 alpha and EF-2 and ribosomal proteins S7 and S10 of the hyperthermophilic archaeum Desulfurococcus mobilis. Mol Gen Genet. 1995 Mar 20;246(6):687–696. [PubMed]
  • Creti R, Ceccarelli E, Bocchetta M, Sanangelantoni AM, Tiboni O, Palm P, Cammarano P. Evolution of translational elongation factor (EF) sequences: reliability of global phylogenies inferred from EF-1 alpha(Tu) and EF-2(G) proteins. Proc Natl Acad Sci U S A. 1994 Apr 12;91(8):3255–3259. [PubMed]
  • DeLong EF. Archaea in coastal marine environments. Proc Natl Acad Sci U S A. 1992 Jun 15;89(12):5685–5689. [PubMed]
  • DeLong EF, Wu KY, Prézelin BB, Jovine RV. High abundance of Archaea in Antarctic marine picoplankton. Nature. 1994 Oct 20;371(6499):695–697. [PubMed]
  • Felsenstein J. Evolutionary trees from DNA sequences: a maximum likelihood approach. J Mol Evol. 1981;17(6):368–376. [PubMed]
  • Fuhrman JA, McCallum K, Davis AA. Novel major archaebacterial group from marine plankton. Nature. 1992 Mar 12;356(6365):148–149. [PubMed]
  • Fuhrman JA, McCallum K, Davis AA. Phylogenetic diversity of subsurface marine microbial communities from the Atlantic and Pacific Oceans. Appl Environ Microbiol. 1993 May;59(5):1294–1302. [PMC free article] [PubMed]
  • Fuller-Pace FV, Nicol SM, Reid AD, Lane DP. DbpA: a DEAD box protein specifically activated by 23s rRNA. EMBO J. 1993 Sep;12(9):3619–3626. [PubMed]
  • Giovannoni SJ, Britschgi TB, Moyer CL, Field KG. Genetic diversity in Sargasso Sea bacterioplankton. Nature. 1990 May 3;345(6270):60–63. [PubMed]
  • Gish W, States DJ. Identification of protein coding regions by database similarity search. Nat Genet. 1993 Mar;3(3):266–272. [PubMed]
  • Hansson M, Rutberg L, Schröder I, Hederstedt L. The Bacillus subtilis hemAXCDBL gene cluster, which encodes enzymes of the biosynthetic pathway from glutamate to uroporphyrinogen III. J Bacteriol. 1991 Apr;173(8):2590–2599. [PMC free article] [PubMed]
  • Iwabe N, Kuma K, Hasegawa M, Osawa S, Miyata T. Evolutionary relationship of archaebacteria, eubacteria, and eukaryotes inferred from phylogenetic trees of duplicated genes. Proc Natl Acad Sci U S A. 1989 Dec;86(23):9355–9359. [PubMed]
  • Kim UJ, Shizuya H, de Jong PJ, Birren B, Simon MI. Stable propagation of cosmid sized human DNA inserts in an F factor based vector. Nucleic Acids Res. 1992 Mar 11;20(5):1083–1085. [PMC free article] [PubMed]
  • Liesack W, Stackebrandt E. Occurrence of novel groups of the domain Bacteria as revealed by analysis of genetic material isolated from an Australian terrestrial environment. J Bacteriol. 1992 Aug;174(15):5072–5078. [PMC free article] [PubMed]
  • Maidak BL, Larsen N, McCaughey MJ, Overbeek R, Olsen GJ, Fogel K, Blandy J, Woese CR. The Ribosomal Database Project. Nucleic Acids Res. 1994 Sep;22(17):3485–3487. [PMC free article] [PubMed]
  • McInerney JO, Wilkinson M, Patching JW, Embley TM, Powell R. Recovery and phylogenetic analysis of novel archaeal rRNA sequences from a deep-sea deposit feeder. Appl Environ Microbiol. 1995 Apr;61(4):1646–1648. [PMC free article] [PubMed]
  • Olsen GJ, Matsuda H, Hagstrom R, Overbeek R. fastDNAmL: a tool for construction of phylogenetic trees of DNA sequences using maximum likelihood. Comput Appl Biosci. 1994 Feb;10(1):41–48. [PubMed]
  • Ouzounis CA, Blencowe BJ. Bacterial DNA replication initiation factor priA is related to proteins belonging to the 'DEAD-box' family. Nucleic Acids Res. 1991 Dec 25;19(24):6953–6953. [PMC free article] [PubMed]
  • Raskin L, Stromley JM, Rittmann BE, Stahl DA. Group-specific 16S rRNA hybridization probes to describe natural communities of methanogens. Appl Environ Microbiol. 1994 Apr;60(4):1232–1240. [PMC free article] [PubMed]
  • Schmidt TM, DeLong EF, Pace NR. Analysis of a marine picoplankton community by 16S rRNA gene cloning and sequencing. J Bacteriol. 1991 Jul;173(14):4371–4378. [PMC free article] [PubMed]
  • Shizuya H, Birren B, Kim UJ, Mancino V, Slepak T, Tachiiri Y, Simon M. Cloning and stable maintenance of 300-kilobase-pair fragments of human DNA in Escherichia coli using an F-factor-based vector. Proc Natl Acad Sci U S A. 1992 Sep 15;89(18):8794–8797. [PubMed]
  • Stahl DA, Lane DJ, Olsen GJ, Pace NR. Analysis of hydrothermal vent-associated symbionts by ribosomal RNA sequences. Science. 1984 Apr 27;224(4647):409–411. [PubMed]
  • Weisburg WG, Tully JG, Rose DL, Petzel JP, Oyaizu H, Yang D, Mandelco L, Sechrest J, Lawrence TG, Van Etten J, et al. A phylogenetic analysis of the mycoplasmas: basis for their classification. J Bacteriol. 1989 Dec;171(12):6455–6467. [PMC free article] [PubMed]
  • Woese CR. Bacterial evolution. Microbiol Rev. 1987 Jun;51(2):221–271. [PMC free article] [PubMed]
  • Woese CR, Achenbach L, Rouviere P, Mandelco L. Archaeal phylogeny: reexamination of the phylogenetic position of Archaeoglobus fulgidus in light of certain composition-induced artifacts. Syst Appl Microbiol. 1991;14(4):364–371. [PubMed]
  • Woese CR, Kandler O, Wheelis ML. Towards a natural system of organisms: proposal for the domains Archaea, Bacteria, and Eucarya. Proc Natl Acad Sci U S A. 1990 Jun;87(12):4576–4579. [PubMed]
  • Zharkikh A, Li WH. Statistical properties of bootstrap estimation of phylogenetic variability from nucleotide sequences. I. Four taxa with a molecular clock. Mol Biol Evol. 1992 Nov;9(6):1119–1147. [PubMed]

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