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Polymerase chain reaction amplification of naphthalene-catabolic and 16S rRNA gene sequences from indigenous sediment bacteria
We report the amplification of bacterial genes from uninoculated surface and subsurface sediments by the polymerase chain reaction (PCR). PCR amplification of indigenous bacterial 16S ribosomal DNA genes was unsuccessful when subsurface sediment containing approximately 10(7) cells.g-1 was added dir...
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| Published in: | Applied and Environmental Microbiology 1993-03, Vol.59 (3), p.687-694 |
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| container_end_page | 694 |
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| container_title | Applied and Environmental Microbiology |
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| creator | Herrick, J.B. (Cornell University, Ithaca, NY) Madsen, E.L Batt, C.A Ghiorse, W.C |
| description | We report the amplification of bacterial genes from uninoculated surface and subsurface sediments by the polymerase chain reaction (PCR). PCR amplification of indigenous bacterial 16S ribosomal DNA genes was unsuccessful when subsurface sediment containing approximately 10(7) cells.g-1 was added directly to a PCR mixture. However, when 10 mg of sediment was inoculated with approximately 10(5) cells of Pseudomonas putida G7, the nahAc naphthalene dioxygenase gene characteristic of the P. putida G7 NAH7 plasmid was detected by PCR amplification. Southern blotting of the PCR amplification product improved sensitivity to 10(3) to 10(4) cells from samples inoculated with P. putida G7, but controls with no sediment added showed that the PCR was partially inhibited by the sediments. Lysozyme - sodium dodecyl sulfate - freeze-thaw DNA extraction was combined with gel electrophoretic partial purification in the presence of polyvinylpyrrolidone to render DNA from indigenous bacteria in surface or subsurface sediment samples amplifiable by PCR using eubacterial 16S ribosomal DNA primers. The nahAc gene could also be amplified from indigenous bacteria by using nahAc-specific primers when PCR conditions were modified by increasing Taq and primer concentrations. Restriction digests of the nahAc amplification products from surface and subsurface sediments revealed polymorphism relative to P. putida G7. The procedures for DNA extraction, purification, and PCR amplification described here demonstrate that the PCR is a potentially useful tool in studies of function- and taxon-specific DNA from indigenous microbial communities in sediment and groundwater environments |
| doi_str_mv | 10.1128/aem.59.3.687-694.1993 |
| format | article |
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(Cornell University, Ithaca, NY) ; Madsen, E.L ; Batt, C.A ; Ghiorse, W.C</creator><creatorcontrib>Herrick, J.B. (Cornell University, Ithaca, NY) ; Madsen, E.L ; Batt, C.A ; Ghiorse, W.C</creatorcontrib><description>We report the amplification of bacterial genes from uninoculated surface and subsurface sediments by the polymerase chain reaction (PCR). PCR amplification of indigenous bacterial 16S ribosomal DNA genes was unsuccessful when subsurface sediment containing approximately 10(7) cells.g-1 was added directly to a PCR mixture. However, when 10 mg of sediment was inoculated with approximately 10(5) cells of Pseudomonas putida G7, the nahAc naphthalene dioxygenase gene characteristic of the P. putida G7 NAH7 plasmid was detected by PCR amplification. Southern blotting of the PCR amplification product improved sensitivity to 10(3) to 10(4) cells from samples inoculated with P. putida G7, but controls with no sediment added showed that the PCR was partially inhibited by the sediments. Lysozyme - sodium dodecyl sulfate - freeze-thaw DNA extraction was combined with gel electrophoretic partial purification in the presence of polyvinylpyrrolidone to render DNA from indigenous bacteria in surface or subsurface sediment samples amplifiable by PCR using eubacterial 16S ribosomal DNA primers. The nahAc gene could also be amplified from indigenous bacteria by using nahAc-specific primers when PCR conditions were modified by increasing Taq and primer concentrations. Restriction digests of the nahAc amplification products from surface and subsurface sediments revealed polymorphism relative to P. putida G7. The procedures for DNA extraction, purification, and PCR amplification described here demonstrate that the PCR is a potentially useful tool in studies of function- and taxon-specific DNA from indigenous microbial communities in sediment and groundwater environments</description><identifier>ISSN: 0099-2240</identifier><identifier>EISSN: 1098-5336</identifier><identifier>DOI: 10.1128/aem.59.3.687-694.1993</identifier><identifier>PMID: 7683182</identifier><identifier>CODEN: AEMIDF</identifier><language>eng</language><publisher>Washington, DC: American Society for Microbiology</publisher><subject>ADN ; AGUAS SUBTERRANEAS ; Animal, plant and microbial ecology ; Applied ecology ; ARN RIBOSOMAL ; ARN RIBOSOMIAL ; BACTERIA ; Base Sequence ; BIOCHIMIE ; Biological and medical sciences ; Biology ; BIOQUIMICA ; Blotting, Southern ; CAPA FREATICA ; Deoxyribonucleic acid ; DESINFECTANT ; DESINFECTANTES ; Dioxygenases ; DNA ; DNA, Bacterial - chemistry ; DNA, Bacterial - isolation & purification ; DNA, Ribosomal - genetics ; EAU SOUTERRAINE ; Ecotoxicology, biological effects of pollution ; FLORA DEL SUELO ; FLORE DU SOL ; Fundamental and applied biological sciences. Psychology ; GENE ; GENES ; Genes, Bacterial ; Molecular Sequence Data ; Multienzyme Complexes - genetics ; NAPPE SOUTERRAINE ; OXIDORREDUCTASAS ; OXYDOREDUCTASE ; Oxygenases - genetics ; POLLUTION DU SOL ; POLUCION DEL SUELO ; Polymerase Chain Reaction - methods ; Polymorphism, Restriction Fragment Length ; PSEUDOMONAS PUTIDA ; Pseudomonas putida - genetics ; RIBOSOMAS ; RIBOSOME ; RNA, Bacterial - genetics ; RNA, Ribosomal, 16S - genetics ; SEDIMENT ; SEDIMENTO ; Sensitivity and Specificity ; Soil Microbiology ; SOL ANTHROPOGENE ; Techniques ; TIPOS ANTROPOGENICOS DE SUELOS</subject><ispartof>Applied and Environmental Microbiology, 1993-03, Vol.59 (3), p.687-694</ispartof><rights>1993 INIST-CNRS</rights><rights>Copyright American Society for Microbiology Mar 1993</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c5033-17e75ac544377d91d958caae61924f93f2beca0d3a667533ee107527a95d97f73</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC202175/pdf/$$EPDF$$P50$$Gpubmedcentral$$H</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC202175/$$EHTML$$P50$$Gpubmedcentral$$H</linktohtml><link.rule.ids>230,315,733,786,790,891,3207,3208,27957,27958,53827,53829</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=4720966$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/7683182$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Herrick, J.B. (Cornell University, Ithaca, NY)</creatorcontrib><creatorcontrib>Madsen, E.L</creatorcontrib><creatorcontrib>Batt, C.A</creatorcontrib><creatorcontrib>Ghiorse, W.C</creatorcontrib><title>Polymerase chain reaction amplification of naphthalene-catabolic and 16S rRNA gene sequences from indigenous sediment bacteria</title><title>Applied and Environmental Microbiology</title><addtitle>Appl Environ Microbiol</addtitle><description>We report the amplification of bacterial genes from uninoculated surface and subsurface sediments by the polymerase chain reaction (PCR). PCR amplification of indigenous bacterial 16S ribosomal DNA genes was unsuccessful when subsurface sediment containing approximately 10(7) cells.g-1 was added directly to a PCR mixture. However, when 10 mg of sediment was inoculated with approximately 10(5) cells of Pseudomonas putida G7, the nahAc naphthalene dioxygenase gene characteristic of the P. putida G7 NAH7 plasmid was detected by PCR amplification. Southern blotting of the PCR amplification product improved sensitivity to 10(3) to 10(4) cells from samples inoculated with P. putida G7, but controls with no sediment added showed that the PCR was partially inhibited by the sediments. Lysozyme - sodium dodecyl sulfate - freeze-thaw DNA extraction was combined with gel electrophoretic partial purification in the presence of polyvinylpyrrolidone to render DNA from indigenous bacteria in surface or subsurface sediment samples amplifiable by PCR using eubacterial 16S ribosomal DNA primers. The nahAc gene could also be amplified from indigenous bacteria by using nahAc-specific primers when PCR conditions were modified by increasing Taq and primer concentrations. Restriction digests of the nahAc amplification products from surface and subsurface sediments revealed polymorphism relative to P. putida G7. The procedures for DNA extraction, purification, and PCR amplification described here demonstrate that the PCR is a potentially useful tool in studies of function- and taxon-specific DNA from indigenous microbial communities in sediment and groundwater environments</description><subject>ADN</subject><subject>AGUAS SUBTERRANEAS</subject><subject>Animal, plant and microbial ecology</subject><subject>Applied ecology</subject><subject>ARN RIBOSOMAL</subject><subject>ARN RIBOSOMIAL</subject><subject>BACTERIA</subject><subject>Base Sequence</subject><subject>BIOCHIMIE</subject><subject>Biological and medical sciences</subject><subject>Biology</subject><subject>BIOQUIMICA</subject><subject>Blotting, Southern</subject><subject>CAPA FREATICA</subject><subject>Deoxyribonucleic acid</subject><subject>DESINFECTANT</subject><subject>DESINFECTANTES</subject><subject>Dioxygenases</subject><subject>DNA</subject><subject>DNA, Bacterial - chemistry</subject><subject>DNA, Bacterial - isolation & purification</subject><subject>DNA, Ribosomal - genetics</subject><subject>EAU SOUTERRAINE</subject><subject>Ecotoxicology, biological effects of pollution</subject><subject>FLORA DEL SUELO</subject><subject>FLORE DU SOL</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>GENE</subject><subject>GENES</subject><subject>Genes, Bacterial</subject><subject>Molecular Sequence Data</subject><subject>Multienzyme Complexes - genetics</subject><subject>NAPPE SOUTERRAINE</subject><subject>OXIDORREDUCTASAS</subject><subject>OXYDOREDUCTASE</subject><subject>Oxygenases - genetics</subject><subject>POLLUTION DU SOL</subject><subject>POLUCION DEL SUELO</subject><subject>Polymerase Chain Reaction - methods</subject><subject>Polymorphism, Restriction Fragment Length</subject><subject>PSEUDOMONAS PUTIDA</subject><subject>Pseudomonas putida - genetics</subject><subject>RIBOSOMAS</subject><subject>RIBOSOME</subject><subject>RNA, Bacterial - genetics</subject><subject>RNA, Ribosomal, 16S - genetics</subject><subject>SEDIMENT</subject><subject>SEDIMENTO</subject><subject>Sensitivity and Specificity</subject><subject>Soil Microbiology</subject><subject>SOL ANTHROPOGENE</subject><subject>Techniques</subject><subject>TIPOS ANTROPOGENICOS DE SUELOS</subject><issn>0099-2240</issn><issn>1098-5336</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1993</creationdate><recordtype>article</recordtype><recordid>eNqFksuKFDEUhgtRxrb1BYSBIOKuylwqSWXhYhi8waDiOOtwOpXqylCVtEm3Mhuf3dN20zhuXIWc_zu35K-qc0Ybxnj3GvzcSNOIRnW6VqZtmDHiQbVg1HS1FEI9rBaUGlNz3tLH1ZNSbimlLVXdWXWmVSdYxxfVry9pupt9huKJGyFEkj24bUiRwLyZwhAc_LmlgUTYjNsRJh99jVFYpSk4ArEnTF2T_PXTBVmjRor_vvPR-UKGnGYSYh8wnnYFlT7MPm7JCnv4HOBp9WiAqfhnx3NZ3bx7--3yQ331-f3Hy4ur2kkqRM201xKcbFuhdW9Yb2TnALxihreDEQNfeQe0F6CUxt29Z1RLrsHI3uhBi2X15lB3s1vNvnc4Q4bJbnKYId_ZBMHeV2IY7Tr9sJxyhhWX1atjfk64XNnaORTnpwmix8WslppxgZ3_BzLVdkpIhuCLf8DbtMsRHwF7SsOlMPux5QFyOZWS_XCamFG7d4FFF1hprLDoAosusHsXYN753-ueso7fjvrLow7FwTRkiC6UE9ZqTo1SiJEDNob1-DNkb6HM91oi8vyADJAsrDNWubk2LZVUK_EbcFrQRg</recordid><startdate>19930301</startdate><enddate>19930301</enddate><creator>Herrick, J.B. (Cornell University, Ithaca, NY)</creator><creator>Madsen, E.L</creator><creator>Batt, C.A</creator><creator>Ghiorse, W.C</creator><general>American Society for Microbiology</general><scope>FBQ</scope><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QL</scope><scope>7QO</scope><scope>7SN</scope><scope>7SS</scope><scope>7ST</scope><scope>7T7</scope><scope>7TM</scope><scope>7U9</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>H94</scope><scope>M7N</scope><scope>P64</scope><scope>RC3</scope><scope>SOI</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>19930301</creationdate><title>Polymerase chain reaction amplification of naphthalene-catabolic and 16S rRNA gene sequences from indigenous sediment bacteria</title><author>Herrick, J.B. (Cornell University, Ithaca, NY) ; Madsen, E.L ; Batt, C.A ; Ghiorse, W.C</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c5033-17e75ac544377d91d958caae61924f93f2beca0d3a667533ee107527a95d97f73</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1993</creationdate><topic>ADN</topic><topic>AGUAS SUBTERRANEAS</topic><topic>Animal, plant and microbial ecology</topic><topic>Applied ecology</topic><topic>ARN RIBOSOMAL</topic><topic>ARN RIBOSOMIAL</topic><topic>BACTERIA</topic><topic>Base Sequence</topic><topic>BIOCHIMIE</topic><topic>Biological and medical sciences</topic><topic>Biology</topic><topic>BIOQUIMICA</topic><topic>Blotting, Southern</topic><topic>CAPA FREATICA</topic><topic>Deoxyribonucleic acid</topic><topic>DESINFECTANT</topic><topic>DESINFECTANTES</topic><topic>Dioxygenases</topic><topic>DNA</topic><topic>DNA, Bacterial - chemistry</topic><topic>DNA, Bacterial - isolation & purification</topic><topic>DNA, Ribosomal - genetics</topic><topic>EAU SOUTERRAINE</topic><topic>Ecotoxicology, biological effects of pollution</topic><topic>FLORA DEL SUELO</topic><topic>FLORE DU SOL</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>GENE</topic><topic>GENES</topic><topic>Genes, Bacterial</topic><topic>Molecular Sequence Data</topic><topic>Multienzyme Complexes - genetics</topic><topic>NAPPE SOUTERRAINE</topic><topic>OXIDORREDUCTASAS</topic><topic>OXYDOREDUCTASE</topic><topic>Oxygenases - genetics</topic><topic>POLLUTION DU SOL</topic><topic>POLUCION DEL SUELO</topic><topic>Polymerase Chain Reaction - methods</topic><topic>Polymorphism, Restriction Fragment Length</topic><topic>PSEUDOMONAS PUTIDA</topic><topic>Pseudomonas putida - genetics</topic><topic>RIBOSOMAS</topic><topic>RIBOSOME</topic><topic>RNA, Bacterial - genetics</topic><topic>RNA, Ribosomal, 16S - genetics</topic><topic>SEDIMENT</topic><topic>SEDIMENTO</topic><topic>Sensitivity and Specificity</topic><topic>Soil Microbiology</topic><topic>SOL ANTHROPOGENE</topic><topic>Techniques</topic><topic>TIPOS ANTROPOGENICOS DE SUELOS</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Herrick, J.B. (Cornell University, Ithaca, NY)</creatorcontrib><creatorcontrib>Madsen, E.L</creatorcontrib><creatorcontrib>Batt, C.A</creatorcontrib><creatorcontrib>Ghiorse, W.C</creatorcontrib><collection>AGRIS</collection><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Biotechnology Research Abstracts</collection><collection>Ecology Abstracts</collection><collection>Entomology Abstracts (Full archive)</collection><collection>Environment Abstracts</collection><collection>Industrial and Applied Microbiology Abstracts (Microbiology A)</collection><collection>Nucleic Acids Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>Engineering Research Database</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Genetics Abstracts</collection><collection>Environment Abstracts</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Applied and Environmental Microbiology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Herrick, J.B. (Cornell University, Ithaca, NY)</au><au>Madsen, E.L</au><au>Batt, C.A</au><au>Ghiorse, W.C</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Polymerase chain reaction amplification of naphthalene-catabolic and 16S rRNA gene sequences from indigenous sediment bacteria</atitle><jtitle>Applied and Environmental Microbiology</jtitle><addtitle>Appl Environ Microbiol</addtitle><date>1993-03-01</date><risdate>1993</risdate><volume>59</volume><issue>3</issue><spage>687</spage><epage>694</epage><pages>687-694</pages><issn>0099-2240</issn><eissn>1098-5336</eissn><coden>AEMIDF</coden><notes>P34</notes><notes>T01</notes><notes>9405076</notes><notes>ObjectType-Article-2</notes><notes>SourceType-Scholarly Journals-1</notes><notes>ObjectType-Feature-1</notes><notes>content type line 23</notes><notes>ObjectType-Article-1</notes><notes>ObjectType-Feature-2</notes><abstract>We report the amplification of bacterial genes from uninoculated surface and subsurface sediments by the polymerase chain reaction (PCR). PCR amplification of indigenous bacterial 16S ribosomal DNA genes was unsuccessful when subsurface sediment containing approximately 10(7) cells.g-1 was added directly to a PCR mixture. However, when 10 mg of sediment was inoculated with approximately 10(5) cells of Pseudomonas putida G7, the nahAc naphthalene dioxygenase gene characteristic of the P. putida G7 NAH7 plasmid was detected by PCR amplification. Southern blotting of the PCR amplification product improved sensitivity to 10(3) to 10(4) cells from samples inoculated with P. putida G7, but controls with no sediment added showed that the PCR was partially inhibited by the sediments. Lysozyme - sodium dodecyl sulfate - freeze-thaw DNA extraction was combined with gel electrophoretic partial purification in the presence of polyvinylpyrrolidone to render DNA from indigenous bacteria in surface or subsurface sediment samples amplifiable by PCR using eubacterial 16S ribosomal DNA primers. The nahAc gene could also be amplified from indigenous bacteria by using nahAc-specific primers when PCR conditions were modified by increasing Taq and primer concentrations. Restriction digests of the nahAc amplification products from surface and subsurface sediments revealed polymorphism relative to P. putida G7. The procedures for DNA extraction, purification, and PCR amplification described here demonstrate that the PCR is a potentially useful tool in studies of function- and taxon-specific DNA from indigenous microbial communities in sediment and groundwater environments</abstract><cop>Washington, DC</cop><pub>American Society for Microbiology</pub><pmid>7683182</pmid><doi>10.1128/aem.59.3.687-694.1993</doi><tpages>8</tpages><oa>free_for_read</oa></addata></record> |
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| identifier | ISSN: 0099-2240 |
| ispartof | Applied and Environmental Microbiology, 1993-03, Vol.59 (3), p.687-694 |
| issn | 0099-2240 1098-5336 |
| language | eng |
| recordid | cdi_pubmed_primary_7683182 |
| source | PubMed Central（OA）; American Society for Microbiology (ASM) Journals |
| subjects | ADN AGUAS SUBTERRANEAS Animal, plant and microbial ecology Applied ecology ARN RIBOSOMAL ARN RIBOSOMIAL BACTERIA Base Sequence BIOCHIMIE Biological and medical sciences Biology BIOQUIMICA Blotting, Southern CAPA FREATICA Deoxyribonucleic acid DESINFECTANT DESINFECTANTES Dioxygenases DNA DNA, Bacterial - chemistry DNA, Bacterial - isolation & purification DNA, Ribosomal - genetics EAU SOUTERRAINE Ecotoxicology, biological effects of pollution FLORA DEL SUELO FLORE DU SOL Fundamental and applied biological sciences. Psychology GENE GENES Genes, Bacterial Molecular Sequence Data Multienzyme Complexes - genetics NAPPE SOUTERRAINE OXIDORREDUCTASAS OXYDOREDUCTASE Oxygenases - genetics POLLUTION DU SOL POLUCION DEL SUELO Polymerase Chain Reaction - methods Polymorphism, Restriction Fragment Length PSEUDOMONAS PUTIDA Pseudomonas putida - genetics RIBOSOMAS RIBOSOME RNA, Bacterial - genetics RNA, Ribosomal, 16S - genetics SEDIMENT SEDIMENTO Sensitivity and Specificity Soil Microbiology SOL ANTHROPOGENE Techniques TIPOS ANTROPOGENICOS DE SUELOS |
| title | Polymerase chain reaction amplification of naphthalene-catabolic and 16S rRNA gene sequences from indigenous sediment bacteria |
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