Phylogenetic screening of a bacterial, metagenomic library using homing endonuclease restriction and marker insertion
Identifieur interne : 000B88 ( Main/Exploration ); précédent : 000B87; suivant : 000B89Phylogenetic screening of a bacterial, metagenomic library using homing endonuclease restriction and marker insertion
Auteurs : Pui Yi Yung [Australie] ; Catherine Burke [Australie] ; Matt Lewis [États-Unis] ; Suhelen Egan [Australie] ; Staffan Kjelleberg [Australie] ; Torsten Thomas [Australie]Source :
- Nucleic Acids Research [ 0305-1048 ] ; 2009.
Abstract
Metagenomics provides access to the uncultured majority of the microbial world. The approaches employed in this field have, however, had limited success in linking functional genes to the taxonomic or phylogenetic origin of the organism they belong to. Here we present an efficient strategy to recover environmental DNA fragments that contain phylogenetic marker genes from metagenomic libraries. Our method involves the cleavage of 23S ribsosmal RNA (rRNA) genes within pooled library clones by the homing endonuclease I-
Url:
DOI: 10.1093/nar/gkp746
PubMed: 19767618
PubMed Central: 2790883
Affiliations:
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Craig Venter Institute, 9704 Medical Center Drive, Rockville, MD 20850, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>J. Craig Venter Institute, 9704 Medical Center Drive, Rockville, MD 20850</wicri:regionArea><placeName><region type="state">Maryland</region></placeName></affiliation></author><author><name sortKey="Egan, Suhelen" sort="Egan, Suhelen" uniqKey="Egan S" first="Suhelen" last="Egan">Suhelen Egan</name><affiliation wicri:level="1"><nlm:aff wicri:cut=" and" id="AFF1">Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW, Australia</nlm:aff><country xml:lang="fr">Australie</country><wicri:regionArea>Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW</wicri:regionArea><wicri:noRegion>2035 NSW</wicri:noRegion></affiliation></author><author><name sortKey="Kjelleberg, Staffan" sort="Kjelleberg, Staffan" uniqKey="Kjelleberg S" first="Staffan" last="Kjelleberg">Staffan Kjelleberg</name><affiliation wicri:level="1"><nlm:aff wicri:cut=" and" id="AFF1">Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW, Australia</nlm:aff><country xml:lang="fr">Australie</country><wicri:regionArea>Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW</wicri:regionArea><wicri:noRegion>2035 NSW</wicri:noRegion></affiliation></author><author><name sortKey="Thomas, Torsten" sort="Thomas, Torsten" uniqKey="Thomas T" first="Torsten" last="Thomas">Torsten Thomas</name><affiliation wicri:level="1"><nlm:aff wicri:cut=" and" id="AFF1">Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW, Australia</nlm:aff><country xml:lang="fr">Australie</country><wicri:regionArea>Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW</wicri:regionArea><wicri:noRegion>2035 NSW</wicri:noRegion></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">PMC</idno><idno type="pmid">19767618</idno><idno type="pmc">2790883</idno><idno type="url">http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2790883</idno><idno type="RBID">PMC:2790883</idno><idno type="doi">10.1093/nar/gkp746</idno><date when="2009">2009</date><idno type="wicri:Area/Pmc/Corpus">000576</idno><idno type="wicri:Area/Pmc/Curation">000576</idno><idno type="wicri:Area/Pmc/Checkpoint">000623</idno><idno type="wicri:Area/Ncbi/Merge">000104</idno><idno type="wicri:Area/Ncbi/Curation">000104</idno><idno type="wicri:Area/Ncbi/Checkpoint">000104</idno><idno type="wicri:doubleKey">0305-1048:2009:Yung P:phylogenetic:screening:of</idno><idno type="wicri:Area/Main/Merge">000B94</idno><idno type="wicri:Area/Main/Curation">000B88</idno><idno type="wicri:Area/Main/Exploration">000B88</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en" level="a" type="main">Phylogenetic screening of a bacterial, metagenomic library using homing endonuclease restriction and marker insertion</title><author><name sortKey="Yung, Pui Yi" sort="Yung, Pui Yi" uniqKey="Yung P" first="Pui Yi" last="Yung">Pui Yi Yung</name><affiliation wicri:level="1"><nlm:aff wicri:cut=" and" id="AFF1">Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW, Australia</nlm:aff><country xml:lang="fr">Australie</country><wicri:regionArea>Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW</wicri:regionArea><wicri:noRegion>2035 NSW</wicri:noRegion></affiliation></author><author><name sortKey="Burke, Catherine" sort="Burke, Catherine" uniqKey="Burke C" first="Catherine" last="Burke">Catherine Burke</name><affiliation wicri:level="1"><nlm:aff wicri:cut=" and" id="AFF1">Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW, Australia</nlm:aff><country xml:lang="fr">Australie</country><wicri:regionArea>Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW</wicri:regionArea><wicri:noRegion>2035 NSW</wicri:noRegion></affiliation></author><author><name sortKey="Lewis, Matt" sort="Lewis, Matt" uniqKey="Lewis M" first="Matt" last="Lewis">Matt Lewis</name><affiliation wicri:level="2"><nlm:aff id="AFF1">J. Craig Venter Institute, 9704 Medical Center Drive, Rockville, MD 20850, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>J. Craig Venter Institute, 9704 Medical Center Drive, Rockville, MD 20850</wicri:regionArea><placeName><region type="state">Maryland</region></placeName></affiliation></author><author><name sortKey="Egan, Suhelen" sort="Egan, Suhelen" uniqKey="Egan S" first="Suhelen" last="Egan">Suhelen Egan</name><affiliation wicri:level="1"><nlm:aff wicri:cut=" and" id="AFF1">Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW, Australia</nlm:aff><country xml:lang="fr">Australie</country><wicri:regionArea>Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW</wicri:regionArea><wicri:noRegion>2035 NSW</wicri:noRegion></affiliation></author><author><name sortKey="Kjelleberg, Staffan" sort="Kjelleberg, Staffan" uniqKey="Kjelleberg S" first="Staffan" last="Kjelleberg">Staffan Kjelleberg</name><affiliation wicri:level="1"><nlm:aff wicri:cut=" and" id="AFF1">Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW, Australia</nlm:aff><country xml:lang="fr">Australie</country><wicri:regionArea>Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW</wicri:regionArea><wicri:noRegion>2035 NSW</wicri:noRegion></affiliation></author><author><name sortKey="Thomas, Torsten" sort="Thomas, Torsten" uniqKey="Thomas T" first="Torsten" last="Thomas">Torsten Thomas</name><affiliation wicri:level="1"><nlm:aff wicri:cut=" and" id="AFF1">Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW, Australia</nlm:aff><country xml:lang="fr">Australie</country><wicri:regionArea>Centre for Marine Bio-Innovation and School of Biotechnology and Biomolecular Sciences, University of New South Wales, 2035 NSW</wicri:regionArea><wicri:noRegion>2035 NSW</wicri:noRegion></affiliation></author></analytic><series><title level="j">Nucleic Acids Research</title><idno type="ISSN">0305-1048</idno><idno type="eISSN">1362-4962</idno><imprint><date when="2009">2009</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p>Metagenomics provides access to the uncultured majority of the microbial world. The approaches employed in this field have, however, had limited success in linking functional genes to the taxonomic or phylogenetic origin of the organism they belong to. Here we present an efficient strategy to recover environmental DNA fragments that contain phylogenetic marker genes from metagenomic libraries. Our method involves the cleavage of 23S ribsosmal RNA (rRNA) genes within pooled library clones by the homing endonuclease I-<italic>Ceu</italic>I followed by the insertion and selection of an antibiotic resistance cassette. This approach was applied to screen a library of 6500 fosmid clones derived from the microbial community associated with the sponge <italic>Cymbastela concentrica</italic>. Several fosmid clones were recovered after the screen and detailed phylogenetic and taxonomic assignment based on the rRNA gene showed that they belong to previously unknown organisms. In addition, compositional features of these fosmid clones were used to classify and taxonomically assign a dataset of environmental shotgun sequences. Our approach represents a valuable tool for the analysis of rapidly increasing, environmental DNA sequencing information.</p></div></front><back><div1 type="bibliography"><listBibl><biblStruct><analytic><author><name sortKey="Ferrer, M" uniqKey="Ferrer M">M Ferrer</name></author><author><name sortKey="Beloqui, A" uniqKey="Beloqui A">A Beloqui</name></author><author><name sortKey="Timmis, Kn" uniqKey="Timmis K">KN Timmis</name></author><author><name sortKey="Golyshin, Pn" uniqKey="Golyshin P">PN Golyshin</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Handelsman, J" uniqKey="Handelsman J">J Handelsman</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Li, X" uniqKey="Li X">X Li</name></author><author><name sortKey="Qin, L" uniqKey="Qin L">L Qin</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Streit, Wr" uniqKey="Streit W">WR Streit</name></author><author><name sortKey="Schmitz, Ra" 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Grossman</name></author><author><name sortKey="Loiacono, Ka" uniqKey="Loiacono K">KA Loiacono</name></author><author><name sortKey="Lynch, Ba" uniqKey="Lynch B">BA Lynch</name></author><author><name sortKey="Phillips, T" uniqKey="Phillips T">T Phillips</name></author><author><name sortKey="Narula, S" uniqKey="Narula S">S Narula</name></author><author><name sortKey="Sundaramoorthi, R" uniqKey="Sundaramoorthi R">R Sundaramoorthi</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Tringe, S" uniqKey="Tringe S">S Tringe</name></author><author><name sortKey="Hugenholtz, P" uniqKey="Hugenholtz P">P Hugenholtz</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Arnould, S" uniqKey="Arnould S">S Arnould</name></author><author><name sortKey="Chames, P" uniqKey="Chames P">P Chames</name></author><author><name sortKey="Perez, C" uniqKey="Perez C">C Perez</name></author><author><name sortKey="Lacroix, E" uniqKey="Lacroix E">E Lacroix</name></author><author><name sortKey="Duclert, A" uniqKey="Duclert A">A Duclert</name></author><author><name sortKey="Epinat, J" uniqKey="Epinat J">J Epinat</name></author><author><name sortKey="Stricher, F" uniqKey="Stricher F">F Stricher</name></author><author><name sortKey="Petit, A" uniqKey="Petit A">A Petit</name></author><author><name sortKey="Patin, A" uniqKey="Patin A">A Patin</name></author><author><name sortKey="Guillier, S" uniqKey="Guillier S">S Guillier</name></author></analytic></biblStruct></listBibl></div1></back></TEI><affiliations><list><country><li>Australie</li><li>États-Unis</li></country><region><li>Maryland</li></region></list><tree><country name="Australie"><noRegion><name sortKey="Yung, Pui Yi" sort="Yung, Pui Yi" uniqKey="Yung P" first="Pui Yi" last="Yung">Pui Yi Yung</name></noRegion><name sortKey="Burke, Catherine" sort="Burke, Catherine" uniqKey="Burke C" first="Catherine" last="Burke">Catherine Burke</name><name sortKey="Egan, Suhelen" sort="Egan, Suhelen" uniqKey="Egan S" first="Suhelen" last="Egan">Suhelen Egan</name><name sortKey="Kjelleberg, Staffan" sort="Kjelleberg, Staffan" uniqKey="Kjelleberg S" first="Staffan" last="Kjelleberg">Staffan Kjelleberg</name><name sortKey="Thomas, Torsten" sort="Thomas, Torsten" uniqKey="Thomas T" first="Torsten" last="Thomas">Torsten Thomas</name></country><country name="États-Unis"><region name="Maryland"><name sortKey="Lewis, Matt" sort="Lewis, Matt" uniqKey="Lewis M" first="Matt" last="Lewis">Matt Lewis</name></region></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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