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Stein J.L., Marsh T.L., Wu K.Y., Shizuya H., DeLong E.F. Characterization of uncultivated prokaryotes: isolation and analysis of a 40-kilobase-pair genome fragment from a planktonic marine archaeon. J Bacteriol. 178:1996;591-599
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Report of the first BAC libraries constructed from the marine environment. Maximal insert sizes were in excess of 150 kb and the average insert size was above 80 kb. The BAC libraries were screened for rRNA using multiplex PCR. rRNA analysis showed the libraries to be composed of known groups previously identified in marine planktonic bacteria, and representatives from uncultivated abundant marine groups, like the SAR86, SAR116 and Roseobacter. The full sequence and first insight into a 60 kb genomic fragment from a planktonic group II Archaea was reported.
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Béjà O., Suzuki M.T., Koonin E.V., Aravind L., Hadd A., Nguyen L.P., Villacorta R., Amjadi M., Garrigues C., Jovanovich S.B., et al. Construction and analysis of bacterial artificial chromosome libraries from a marine microbial assemblage. Environ Microbiol. 2:2000;516-529 Report of the first BAC libraries constructed from the marine environment. Maximal insert sizes were in excess of 150 kb and the average insert size was above 80 kb. The BAC libraries were screened for rRNA using multiplex PCR. rRNA analysis showed the libraries to be composed of known groups previously identified in marine planktonic bacteria, and representatives from uncultivated abundant marine groups, like the SAR86, SAR116 and Roseobacter. The full sequence and first insight into a 60 kb genomic fragment from a planktonic group II Archaea was reported.
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Cloning the soil metagenome: A strategy for accessing the genetic and functional diversity of uncultured microorganisms
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The authors report the construction and functional screening of the first metagenomic libraries from soil with an average insert size of 27 and 44.5 kb and containing a total of 1 Gb of DNA. Several functional genes were shown to be expressed from the BACs by phenotypic screening.
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Rondon M.R., August P.R., Bettermann A.D., Brady S.F., Grossman T.H., Liles M.R., Loiacono K.A., Lynch B.A., MacNeil I.A., Minor C., et al. Cloning the soil metagenome: a strategy for accessing the genetic and functional diversity of uncultured microorganisms. Appl Environ Microbiol. 66:2000;2541-2547 The authors report the construction and functional screening of the first metagenomic libraries from soil with an average insert size of 27 and 44.5 kb and containing a total of 1 Gb of DNA. Several functional genes were shown to be expressed from the BACs by phenotypic screening.
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Preston CM, Suzuki MT, Béjà O, de la Torre RJ, Steward GF, DeLong ED: Quantitative phylogenetic screening of ribosomal RNA gene-containing clones in Bacterial Artificial Chromosome (BAC) libraries from different depths in Monterey Bay. Microb Ecol: in press.
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Preston, C.M.1
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The first example of the use of the ecogenomics approach to understand the biochemistry and physiology of widespread uncultivated bacterial groups. Specifically, a 130 kb genome fragment from an uncultured marine bacterium of the SAR86 group (γ-proteobacterial group) was isolated on a BAC clone. Shotgun sequencing of the fragment revealed a new type of rhodopsin (proteorhodopsin) related to the archaeal bacteriorhodopsins. Expression and biochemical characterization showed that the protein functions as a light-driven proton pump, thus identifying the first bacterial rhodopsin and a potentially new type of light-harvesting mechanism that appears to be widespread in the oceans.
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Béjà O., Aravind L., Koonin E.V., Suzuki M.T., Hadd A., Nguyen L.P., Jovanovich S.B., Gates C.M., Feldman R.A., Spudich J.L., et al. Bacterial rhodopsin: evidence for a new type of phototrophy in the sea. Science. 289:2000;1902-1906 The first example of the use of the ecogenomics approach to understand the biochemistry and physiology of widespread uncultivated bacterial groups. Specifically, a 130 kb genome fragment from an uncultured marine bacterium of the SAR86 group (γ-proteobacterial group) was isolated on a BAC clone. Shotgun sequencing of the fragment revealed a new type of rhodopsin (proteorhodopsin) related to the archaeal bacteriorhodopsins. Expression and biochemical characterization showed that the protein functions as a light-driven proton pump, thus identifying the first bacterial rhodopsin and a potentially new type of light-harvesting mechanism that appears to be widespread in the oceans.
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Science
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Béjà, O.1
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Béjà O., Spudich E.N., Spudich J.L., Leclerc M., DeLong E.F. Proteorhodopsin phototrophy in the ocean. Nature. 411:2001;786-789
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The authors used hybridization methods to screen marine BAC libraries for the presence of proteorhodopsin. This study is the first to report evidence for a proteorhodopsin in the genome of a member of the α-proteobacterial group.
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de la Torre R.J., Christianson L., Béjà O., Suzuki M.T., Karl D., Heidelberg J.F., DeLong E.F. Proteorhodopsin genes are widely distributed among divergent bacterial taxa. Proc Natl Acad Sci USA. 100:2003;12830-12835 The authors used hybridization methods to screen marine BAC libraries for the presence of proteorhodopsin. This study is the first to report evidence for a proteorhodopsin in the genome of a member of the α-proteobacterial group.
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Man D., Wang W., Sabehi G., Aravind L., Post A.F., Massana R., Spudich E.N., Spudich J.L., Béjà O. Diversification and spectral tuning in marine proteorhodopsins. EMBO J. 22:2003;1725-1731
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Sabehi G., Massana R., Bielawski J.P., Rosenberg M., DeLong E.F., Béjà O. Novel proteorhodopsin variants from the Mediterranean and Red Seas. Environ Microbiol. 5:2003;842-849
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Kolber Z.S., Plumley F.G., Lang A.S., Beatty J.T., Blankenship R.E., VanDover C.L., Vetriani C., Koblízek M., Rathgeber C., Falkowski P.G. Contribution of aerobic photoheterotrophic bacteria to the carbon cycle in the ocean. Science. 292:2001;2492-2495
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Describes the identification of uncultured bacterial groups involved in oceanic aerobic anoxygenic photosynthesis. This paper combines the screening of environmental BAC libraries for the presence of pufML-containing clones using multiplex PCR and comparative genomics.
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Béjà O., Suzuki M.T., Heidelberg J.F., Nelson W.C., Preston C.M., Hamada T., Eisen J.A., Fraser C.M., DeLong E.F. Unsuspected diversity among marine aerobic anoxygenic phototrophs. Nature. 415:2002;630-633 Describes the identification of uncultured bacterial groups involved in oceanic aerobic anoxygenic photosynthesis. This paper combines the screening of environmental BAC libraries for the presence of pufML-containing clones using multiplex PCR and comparative genomics.
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Clone libraries targeting a conserved region of the α subunit of methyl coenzyme M reductase (encoded by the mcrA gene), the enzyme catalyzing the terminal step in methanogenesis, were generated from methane-rich sediments in an attempt to identify enzymes involved in anaerobic methane oxidation. The data was compared with results coming directly from environmental samples, laboratory-incubated microcosms and fosmid libraries.
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Hallam S.J., Girguis P.R., Preston C.M., Richardson P.M., DeLong E.F. Identification of methyl coenzyme M reductase A (mcrA) genes associated with methane-oxidizing Archaea. Appl Environ Microbiol. 69:2003;5483-5491 Clone libraries targeting a conserved region of the α subunit of methyl coenzyme M reductase (encoded by the mcrA gene), the enzyme catalyzing the terminal step in methanogenesis, were generated from methane-rich sediments in an attempt to identify enzymes involved in anaerobic methane oxidation. The data was compared with results coming directly from environmental samples, laboratory-incubated microcosms and fosmid libraries.
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A combination of hardcore biochemistry and marine ecogenomics (using fosmid libraries) to find the enzymes involved in anaerobic methane oxidation in marine sediments.
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