Chapter 86 : Metagenomic Methods for the Identification of Active Microorganisms and Genes in Biodegradation Processes

MyBook is a cheap paperback edition of the original book and will be sold at uniform, low price.

Preview this chapter:
Zoom in

Metagenomic Methods for the Identification of Active Microorganisms and Genes in Biodegradation Processes, Page 1 of 2

| /docserver/preview/fulltext/10.1128/9781555815882/9781555813796_Chap86-1.gif /docserver/preview/fulltext/10.1128/9781555815882/9781555813796_Chap86-2.gif


Isolated microorganisms constitute only a minor fraction of the global microbial diversity, which may comprise millions of species. This chapter discusses some cautionary guidelines that should be followed in the attempts to link phylogeny with function. The simplest and most commonly used approach to identify organisms involved in biodegradation is to isolate microbial strains capable of utilizing the target substrate, xenobiotics, or pollutant as a sole C/N source. Sequence-dependent approaches to identify genes are largely limited by an a priori knowledge of gene(s) mediating the biodegradation processes but have proven invaluable for an understanding of the key enzymes in these metabolic pathways. The majority of control in prokaryotes is thought to occur at the transcriptional level, so the presence of an mRNA is strong evidence for the expression of that gene product. Methods for amplifying genes requiring only one gene-specific primer impose less sequence-dependent bias than standard two-primer PCR amplification procedures. These PCR-based strategies have been used for the recovery of the up- or downstream regions flanking a single PCR primer for the recovery of full-length genes. The minimum number of clones that need to be screened in order to find a gene of interest in the metagenomic library increases with the presence of eukaryotic DNA (a eukaryotic genome is 3 to 140,000 Mbp compared to prokaryotic organisms of 0.6 to 9.5 Mbp). In summary, metagenomic techniques have begun to address both the identity of active organisms and their gene products that mediate biodegradation.

Citation: Cowan D, Stafford W. 2007. Metagenomic Methods for the Identification of Active Microorganisms and Genes in Biodegradation Processes, p 1079-1088. In Hurst C, Crawford R, Garland J, Lipson D, Mills A, Stetzenbach L (ed), Manual of Environmental Microbiology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555815882.ch86

Key Concept Ranking

Denaturing Gradient Gel Electrophoresis
Highlighted Text: Show | Hide
Loading full text...

Full text loading...


1. Akopyants, N. S.,, A. Fradkov,, L. Diatchenko,, J. E. Hill,, P. D. Siebert,, S. A. Lukyanov,, E. D. Sverdlov, and, D. E. Berg. 1998. PCR-based subtractive hybridization and differences in gene content among strains of Helicobacter pylori. Proc. Natl. Acad. Sci. USA 95:1310813113.
2. Amann, R. I.,, B. J. Binder,, R. J. Olson,, S. W. Chisholm,, R. Devereux, and, D. A. Stahl. 1990. Combination of 16S rRNA-targeted oligonucleotide probes with flow cytometry for analyzing mixed microbial populations. Appl. Environ. Microbiol. 56:19191925.
3. Amann, R. I.,, L. Krumholz, and, D. A. Stahl. 1990. Fluorescent oligonucleotide probing of whole cells for determinative, phylogenetic, and environmental studies in microbiology. J. Bacteriol. 172:762770.
4. Amman, R. L.,, W. Ludwig, and, K. Schleifer. 1995. Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol. Rev. 59:143169.
5. Artursson, V.,, and J. K. Jansson. 2003. Use of bromodeoxyuridine immunocapture to identify active bacteria associated with arbuscular mycorrhizal hyphae. Appl. Environ. Microbiol. 69:62086215.
6. Bart, A.,, J. Dankert, and, A. van der Ende. 2000. Representational difference analysis of Neisseria meningitidis identifies sequences that are specific for the hyper-virulent lineage III clone. FEMS Microbiol. Lett. 188:111114.
7. Becker, P.,, W. Hufnagle,, G. Peters, and, M. Herrmann. 2001. Detection of differential gene expression in biofilm-forming versus planktonic populations of Staphylococcus aureus using micro-representational-difference analysis. Appl. Environ. Microbiol. 67:29582965.
8. Beja, O. 2004. To BAC or not to BAC: marine ecogenomics. Curr. Opin. Biotechnol. 15:187190.
9. Beja, O.,, M. T. Suzuki,, E. V. Koonin,, L. Aravind,, A. Hadd,, L. P. Nguyen,, R. Villacorta,, M. Amjadi,, C. Garrigues,, S. B. Jovanovich,, R. A. Feldman, and, E. F. DeLong. 2000. Construction and analysis of bacterial artificial chromosome libraries from a marine microbial assemblage. Environ. Microbiol. 2:516529.
10. Bell, P.,, A. Sunna,, M. D.Gibbs,, N. C. Curach,, H. Nevalainen, and, P. L. Bergquist. 2002. Prospecting for novel lipase genes using PCR. Microbiology 148:22832291.
11. Bohannan, B. J. M.,, and J. Hughes. 2003. New approaches to analyzing microbial biodiversity data. Curr. Opin. Microbiol. 6:282287.
12. Bonin, P. C.,, V. D. Michotey,, A. Mouzdahir, and, J.-F. Rontani. 2002. Anaerobic biodegradation of squalene: using DGGE to monitor the isolation of denitrifying bacteria taken from enrichment cultures. FEMS Microbiol. Ecol. 42:3749.
13. Borneman, J. 1999. Culture-independent identification of microorganisms that respond to specified stimuli. Appl. Environ. Microbiol. 65:33983400.
14. Bowler, L. D.,, M. Hubank, and, B. G. Spratt. 1999. Representational difference analysis of cDNA for the detection of differential gene expression in bacteria: development using a model of iron-regulated gene expression in Neisseria meningitidis. Microbiology 145:35293537.
15. Braker, G.,, A. Fesefeldt, and, K.-P. Witzel. 1998. Development of PCR primer systems for amplification of nitrite reductase genes (nirK and nirS) to detect denitrifying bacteria in environmental samples. Appl. Environ. Microbiol. 64:37693775.
16. Breitbart, M.,, P. Salamon,, B. Andresen,, J. M. Mahaffy,, A. M. Segall,, D. Mead,, F. Azam, and, F. Rohwer. 2002. Genomic analysis of uncultured marine viral communities. Proc. Natl. Acad. Sci. USA 99:1425014255.
17. Cases, I.,, and V. de Lorenzo. 2005. Promoters in the environment: transcription regulation in its natural context. Nat. Microbiol. Rev. 3:105118.
18. Clarke, L.,, and J. Carbon. 1976. A colony bank containing synthetic ColE1 hybrid plasmids representative of the entire E. coli genome. Cell 9:91100.
19. Corey, D. R. 1997. Peptide nucleic acids: expanding the scope of nucleic acid recognition. Trends Biotechnol. 15:224229.
20. Courtois, S.,, C. M. Cappellano,, M. Ball,, F. X. Francou,, P. Normand,, G. Helynck,, A. Martinez,, S. J. Kolvek,, J. Hopke,, M. S. Osburne,, P. R. August,, R. Nalin,, M. Guerineau,, P. Jeannin,, P. Simonet, and, J. L. Pernodet. 2003. Recombinant environmental libraries provide access to microbial diversity for drug discovery from natural products. Appl. Environ. Microbiol. 69:4955.
21. Cowan, D. A. 2000. Microbial genomes—the untapped resource. Trends Biotechnol. 18:1416.
22. Curtis, T. P.,, W. T. Sloan, and, J. W. Scannell. 2002. Estimating prokaryotic diversity and its limits. Proc. Natl. Acad. Sci. USA 99:1049410499.
23. DeLong, E. F.,, L. T. Taylor,, T. L. Marsh, and, C. M. Preston. 1999. Visualization and enumeration of marine planctonic archaea and bacteria by using polyribonucleotide probes and fluorescent in situ hybridization. Appl. Environ. Microbiol. 65:55545563.
24. Diaz, E. 2004. Bacterial degradation of aromatic pollutants: a paradigm of metabolic versatility. Int. Microbiol. 7:173180.
25. Eastwood, R. F.,, E. S. Lagudah, and, R. Appels. 1994. A directed search for DNA sequences tightly linked to cereal cyst nematode resistance in Triticum tauschii. Genome 37:311319.
26. Frischer, M. E.,, J. M. Danforth,, M. A. Newton Healy, and, F. M. Saunders. 2000. Whole-cell versus total RNA extraction for analysis of microbial community structure with 16S rRNA-targeted oligonucleotide probes in salt marsh sediments. Appl. Environ. Microbiol. 66:30373043.
27. Fuchs, B. M.,, G. Wallner,, W. Beisker,, I. Schwippl,, W. Ludwig, and, R. Amann. 1998. Flow cytometric analysis of the in situ accessibility of Escherichia coli 16S rRNA for fluorescently labeled oligonucleotide probes. Appl. Environ. Microbiol. 64:49734982.
28. Futamata, H.,, S. Harayama, and, K. Watanabe. 2001. Group-specific monitoring of phenol hydroxylase genes for a functional assessment of phenol-stimulated trichloroethylene bioremediation. Appl. Environ. Microbiol. 67:46714677.
29. Gabor, E. M.,, W. B. L. Alkema, and, D. B. Janssen. 2004. Quantifying the accessibility of the metagenome by random expression cloning techniques. Environ. Microbiol. 6:879886.
30. Galbraith, E. A.,, D. A. Antonopoulos, and, B. A. White. 2004. Suppressive subtractive hybridisation as a tool for identifying genetic diversity in an environmental metagenome: the rumen as a model. Environ. Microbiol. 6:928937.
31. Gelsomino, A.,, A. C. Keyzer-Wolters,, G. Cacco, and, J. D. van Elsas. 1999. Assessment of bacterial community structure in soil by polymerase chain reaction and denaturing gradient gel electrophoresis. J. Microbiol. Methods 38:15.
32. Ginige, M. P.,, P. Hugenholtz,, H. Daims,, M. Wagner,, J. Keller, and, L. L. Blackall. 2004. Use of stable-isotope probing, full-cycle rRNA analysis, and fluorescence in situ hybridization-microautoradiography to study a methanolfed denitrifying microbial community. Appl. Environ. Micro-biol. 70:588596.
33. Ginzinger, D. G. 2002. Gene quantification using real-time quantitative PCR: an emerging technology hits the mainstream. Exp. Hematol. 30:503512.
34. Green, C. D.,, J. F. Simons,, B. E. Taillon, and, D. A. Lewin. 2001. Open systems: panoramic views of gene expression. J. Immunol. Methods 250:6779.
35. Griffiths, R. I.,, A. S. Whiteley,, A. G. O’Donnell, and, M. J. Bailey. 2000. Rapid method for coextraction of DNA and RNA from natural environments for analysis of ribosomal DNA- and rRNA-based microbial community composition. Appl. Environ. Microbiol. 66:54885491.
36. Habe, H.,, and T. Omori. 2003. Genetics of polycyclic aromatic hydrocarbon metabolism in diverse aerobic bacteria. Biosci. Biotechnol. Biochem. 67:225243.
37. Hallin, S.,, and P. Lindgren. 1999. PCR detection of genes encoding nitrite reductase in denitrifying bacteria. Appl. Environ. Microbiol. 65:16521657.
38. Handelsman, J.,, M. R. Rondon,, S. F. Brady,, J. Clardy, and, R. M. Goodman. 1998. Molecular biological access to the chemistry of unknown soil microbes: a new frontier for natural products. Chem. Biol. 5:R245R249.
39. Henckel, T.,, U. Jackel,, S. Schnell, and, R. Conrad. 2000. Molecular analyses of novel methanotrophic communities in forest soil that oxidize atmospheric methane. Appl. Environ. Microbiol. 66:18011808.
40. Hugenholtz, P.,, B. M. Goebel, and, N. R. Pace. 1998. Impact of culture-independent studies on the emerging phylogenetic view of bacterial diversity. J. Bacteriol. 180:47654774.
41. Hurt, R. A. 2001. Simultaneous recovery of RNA and DNA from soils and sediments. Appl. Environ. Microbiol. 67:44954503.
42. Jeon, C. O.,, W. Park,, P. Padmanabhan,, C. DeRito,, J. R. Snape, and, E. L. Madsen. 2003. Discovery of a bacterium, with distinctive dioxygenase, that is responsible for in situ biodegradation in contaminated sediment. Proc. Nat. Acad. Sci. USA 100:1359113596.
43. Kaeberlein, T.,, K. Lewis, and, S. S. Epstein. 2005. Isolating “uncultivable” microorganisms in pure culture using a simulated natural environment. Science 296:11271129
44. Kirk, J. L.,, L. A. Beaudette,, M. Hart,, P. Moutoglis,, J. N. Klironomos,, H. Lee, and, J. T. Trevors. 2004. Methods of studying soil microbial diversity. J. Microbiol. Methods 58:169188.
45. Knietsch, A.,, T. Waschkowitz,, S. Bowien,, A. Henne, and, R. Daniel. 2003. Metagenomes of complex microbial consortia derived from different soils as sources for novel genes conferring formation of carbonyls from short-chain polyols on Escherichia coli. J. Mol. Microbiol. Biotechnol. 5:4656.
46. Lee, N.,, P. H. Nielsen,, K. H. Andreasen,, S. Juretschko,, J. L. Nielsen,, K.-H. Schleifer, and, M. Wagner. 1999. Combination of fluorescent in situ hybridization and microautoradiography—a new tool for structure-function analyses in microbial ecology. Appl. Environ. Microbiol. 65:12891297.
47. Liu, W.-T.,, and D. A. Stahl. 2002. Molecular approaches for the measurement of density, diversity, and phylogeny, p. 114–134. In C. J. Hurst,, R. L. Crawford,, G. R. Knudsen,, M. J. McInerney, and, L. D. Stetzenbach (ed.), Manual of Environmental Microbiology, 2nd ed. ASM Press, Washington, D.C.
48. Liu, Y.,, and R. F. Whittier. 1995. Thermal asymmetric interlaced PCR: automatable amplification and sequencing of insert end fragments from pi and yac clones for chromosome walking. Genomics 25:674681.
49. Lorenz, P.,, and C. Schleper. 2002. Metagenome—a challenging source of enzyme discovery. J. Mol. Catal. B 19:1319.
50. Loy, A.,, and M. W. M. Horn. 2003. probeBase—an online resource for rRNA-targeted oligonucleotide probes. Nucleic Acids Res. 31:514516.
51. Loy, A. A.,, N. Lehner,, J. Lee,, H. Adamczyk,, J. Meier,, H. Schleifer, and, M. Wagner. 2002. Oligonucleotide microarray for 16S rRNa-based detections of all recognized lineages of sulphate-reducing prokaryotes in the environment. Appl. Environ. Microbiol. 68:50645081.
52. Lukyanov, S. A.,, N. G. Gurskaya,, K. A. Lukyanov,, V. S. Tarabykin, and, E. D. Sverdlov. 1994. Highly efficient subtractive hybridisation of cDNA. Bioorg. Khim. 20:701704.
53. MacGregor, B. J.,, V. Brüchert,, S. Fleischer, and, R. Amann. 2002. Isolation of small-subunit rRNA for stable-isotopic characterization. Environ. Microbiol. 4:451464.
54. MacNaughton, S. J.,, J. R. Stephen,, A. D. Venosa,, G. A. Davis,, Y. J. Chang, and, D. C. White. 1999. Microbial population changes during bioremediation of an experimental oil spill. Appl. Environ. Microbiol. 65:35663574.
55. Manefield, M.,, A. S. Whiteley,, R. I. Griffiths, and, M. J. Bailey. 2002. RNA stable isotope probing, a novel means of linking microbial community function to phylogeny. Appl. Environ. Microbiol. 68:53675373.
56. Martinez, A.,, S. J. Kolvek,, C. L. T. Yip,, J. Hopke,, K. A. Brown,, I. A. MacNeil, and, M. S. Osburne. 2004. Genetically modified bacterial strains and novel bacterial artificial chromosome shuttle vectors for constructing environmental libraries and detecting heterologous natural products in multiple expression hosts. Appl. Environ. Microbiol. 70:24522463.
57. McDonald, I. R.,, E. M. Kenna, and, J. C. Murrell. 1995. Detection of methanotrophic bacteria in environmental samples with the PCR. Appl. Environ. Microbiol. 61:116121.
58. McRae, B. M.,, T. M. LaPara, and, R. M. Hozalski. 2004. Biodegradation of haloacetic acids by bacterial enrichment cultures. Chemosphere 55:915925.
59. Megonigal, M. D.,, E. F. Rappaport,, R. B. Wilson,, D. H. Jones,, J. A. Whitlock,, J. A. Ortega,, D. J. Slater,, P. C. Nowell, and, C. A. Felix. 2000. Panhandle PCR for cDNA: a rapid method for isolation of MLL fusion transcripts involving unknown partner genes. Proc. Natl. Acad. Sci. USA 97:95979602.
60. Mesarch, M. B.,, C. H. Nakatsu, and, L. Nies. 2000. Development of catechol 2,3-dioxygenase-specific primers for monitoring bioremediation by competitive quantitative PCR. Appl. Environ. Microbiol. 66:678683.
61. Metcalfe, A. C.,, M. Krsek,, G. W. Gooday,, J. I. Prosser, and, E. M. H. Wellington. 2002. Molecular analysis of a bacterial chitinolytic community in an upland pasture. Appl. Environ. Microbiol. 68:50425050.
62. Mishra, R. N.,, S. L. Singla-Pareek,, S. Nair,, S. K. Sopory, and, M. K. Reddy. 2002. Directional genome walking using PCR. BioTechniques 33:830834.
63. Moter, A.,, and U. B. Globel. 2000. Fluorescence in situ hybridization (FISH) for direct visualization of microorganisms. J. Microbiol. Methods 41:85112.
64. Moyer, C. L.,, J. M. Tiedje,, F. C. Dobbs, and, D. M. Karl. 1996. A computer-simulated restriction fragment length polymorphism analysis of bacterial small-subunit rRNA genes: efficacy of selected tetrameric restriction enzymes for studies of microbial diversity in nature. Appl. Environ. Microbiol. 62:25012507.
65. Myers, R. M.,, S. G. Fisher,, L. S. Lerman, and, T. Maniatis. 1985. Nearly all single base substitutions in DNA fragments joined to a GC-clamp can be detected by denaturing gradient gel electrophoresis. Nucleic Acids Res. 13:31313145.
66. Myrick, K. V.,, and W. M. Gelbart. 2002. Universal fast walking for direct and versatile determination of flanking sequence. Gene 284:125131.
67. Nicolaisen, M. H.,, and N. B. Ramsing. 2002. Denaturing gradient gel electrophoresis (DGGE) approaches to study the diversity of ammonia-oxidizing bacteria. J. Microbiol. Methods 50:189203.
68. Ochman, H.,, F. J. Ayala, and, D. L. Hartl. 1993. Use of polymerase chain reaction to amplify segments outside boundaries of known sequences. Methods Enzymol. 218:309321.
69. Oliver, J.,, L. Nilsson, and, S. Kjelleberg. 1991. Formation of nonculturable Vibrio vulnificus cells and its relationship to the starvation state. Appl. Environ. Microbiol. 57:26402644.
70. Orphan, V. J.,, C. H. House,, K. U. Hinrichs,, K. D. McKeegan, and, E. F. DeLong. 2002. Multiple archaeal groups mediate methane oxidation in anoxic cold seep sediments. Proc. Natl. Acad. Sci. USA 99:76637668.
71. Paez, J. G.,, M. Lin,, R. Beroukhim,, J. C. Lee,, X. Zhao,, D. J. Richter,, S. Gabriel,, P. Herman,, H. Sasaki,, D. Altshuler,, C. Li,, M. Meyerson, and, W. R. Sellers. 2004. Genome coverage and sequence fidelity of ϕ29 polymerase-based multiple strand displacement whole genome amplification. Nucleic Acids Res. 32:e71.
72. Pearson, A.,, A. L. Sessions,, K. J. Edwards, and, J. M. Meyers. 2004. Phylogenetically specific separation of rRNA from prokaryotes for isotopic analysis. Mar. Chem. 92:295306.
73. Pelz, O.,, A. Chatzinotas,, N. Andersen,, S. M. Bernasconi,, C. Hesse,, W. R. Abraham, and, J. Zeyer. 2001. Use of isotopic and molecular techniques to link toluene degradation in denitrifying aquifer microcosms to specific microbial populations. Arch. Microbiol. 175:270281.
74. Pernthaler, A.,, and R. Amann. 2004. Simultaneous fluorescence in situ hybridization of mRNA and rRNA in environmental bacteria. Appl. Environ. Microbiol. 70:54265433.
75. Prescott, A. M.,, and C. R. Fricker. 1999. Use of PNA oligonucleotides for the in situ detection of Escherichia coli in water samples. Mol. Cell. Probes 13:261268.
76. Purkhold, U.,, M. Wagner,, G. Timmermann,, A. Pommerening-Roser, and, H. P. Koops. 2003. 16S rRNA and amoA-based phylogeny of 12 novel betaproteobacterial ammonia-oxidizing isolates: extension of the dataset and proposal of a new lineage within the nitrosomonads. Int. J. Syst. Evol. Microbiol. 53:14851494.
77. Radajewski, S.,, I. R. McDonald, and, J. C. Murrell. 2003. Stable-isotope probing of nucleic acids: a window to the function of uncultured microorganisms. Curr. Opin. Biotechnol. 14:296302.
78. Radajewski, S.,, G. Webster,, D. S. Reay,, S. A. Morris,, P. Ineson,, D. B. Nedwell,, J. I. Prosser, and, J. C. Murrell. 2002. Identification of active methylotroph populations in an acidic forest soil by stable-isotope probing. Microbiology 148:23312342.
79. Rees, H. C.,, W. D. Grant,, B. E. Jones, and, S. Heaphy. 2004. Diversity of Kenyan soda lake alkaliphiles assessed by molecular methods. Extremophiles 8:6371.
80. Rhee, S.-K.,, X. Liu,, L. Wu,, S. C. Chong,, X. Wan, and, J. Zhou. 2004. Detection of genes involved in the biodegradation and biotransformation in microbial communities by using 50-mer oligonucleotide arrays. Appl. Environ. Microbiol. 70:43034317.
81. Roling, W. F.,, I. R. de Brito Couto,, R. P. Swannell, and, I. M. Head. 2004. Response of archaeal communities in beach sediments to spilled oil and bioremediation. Appl. Environ. Microbiol. 70:26142620.
82. Roling W. F.,, M. G. Milner,, D. M. Jones,, F. Fratepietro,, R. P. Swannell,, F. Daniel, and, I. M. Head. 2004. Bacterial community dynamics and hydrocarbon degradation during a field-scale evaluation of bioremediation on a mudflat beach contaminated with buried oil. Appl. Environ. Micro-biol. 70:26032613.
83. Rondon, M. R.,, P. R. August,, A. D. Bettermann,, S. F. Brady,, T. H. Grossman,, M. R. Liles,, K. A. Loiacono,, B. A. Lynch,, I. A. Macneil,, C. Minor,, C. L. Tiong,, M. Gilman,, M. S. Osburne,, J. Clardy,, J. Handelsman, and, R. M. Goodman. 2000. Cloning the soil metagenome: a strategy for accessing the genetic and functional diversity of uncultured microorganisms. Appl. Environ. Microbiol. 66:25412547.
84. Rose, T. M.,, E. R. Schultz,, J. G. Henikoff,, S. Pietrokovski,, C. M. McCallum, and, S. Henikoff. 1998. Consensus-degenerate hybrid oligonucleotide primers for amplification of distantly related sequences. Nucleic Acids Res. 26:16281635.
85. Roszak, D. B.,, D. J. Grimes, and, R. R. Colwell. 1984. Viable but nonrecoverable stage of Salmonella enteritidis in aquatic systems. Can. J. Microbiol. 30:334338.
86. Schloss, P. D.,, and J. Handelsman. 2003. Biotechno-logical prospects from metagenomics. Curr. Opin. Biotechnol. 14:303310.
87. Schönhuber, W.,, B. Fuchs,, S. Juretschko, and, R. Amann. 1997. Improved sensitivity of whole-cell hybridization by the combination of horseradish peroxidase-labeled oligonucleotides and tyramide signal amplification. Appl. Environ. Microbiol. 63:32683273.
88. Sebat, J. L.,, F. S. Colwell, and, R. L. Crawford. 2003. Metagenomic profiling: microarray analysis of a metagenomic DNA library. Appl. Environ. Microbiol. 69:49274934.
89. Sheffield, V. C.,, D. R. Cox,, L. S. Lerman, and, R. M. Myers. 1989. Attachment of a 40-base-pair GC rich sequence (GC-clamp) to genomic DNA fragments by the polymerase chain-reaction results in improved detection of single-base changes. Proc. Natl. Acad. Sci. USA 86:232236.
90. Sheu, D.,, Y. Wang, and, C. Lee. 2000. Rapid detection of polyhydroxyalkanoate-accumulating bacteria isolated from the environment by colony PCR. Microbiology 146:2019-2025.
91. Short, J. M.,, and E. J. Mathur. December 1999. Production and Use of Normalized DNA Libraries. U.S. patent 6,001,574.
92. Small, J.,, D. R. Call,, F. J. Brockman,, T. M. Straub, and, D. P. Chandler. 2001. Direct detection of 16S rRNA in soil extracts by using oligonucleotide microarrays. Appl. Environ. Microbiol. 67:47084716.
93. Snaidr, J.,, B. Fuchs,, G. Wallner,, M. Wagner,, K. H. Schleifer, and, R. Amann. 1999. Phylogeny and in situ identification of a morphologically conspicuous bacterium, Candidatus Magnospira bakii, present at very low frequency in activated sludge. Environ. Microbiol. 1:125135.
94. Stull, D.,, and J. M. Pisano. 2001. Purely RNA: new innovations enhance the quality, speed, and efficiency of RNA isolation techniques. Scientist 15:2931.
95. Suzuki, M. T.,, and S. J. Giovannoni. 1996. Bias caused by template annealing in the amplification of mixtures of 16S rRNA genes by PCR. Appl. Environ. Microbiol. 62:625630.
96. Thompson, J.,, L. Marcelino, and, M. F. Polz. 2002. Heteroduplexes in mixed-template amplifications: formation, consequence and elimination by ‘reconditioning PCR.’ Nucleic Acids Res. 30:20832088.
97. Torsvik, V.,, and L. Ovreas. 2002. Microbial diversity and function in soil: from genes to ecosystems. Curr. Opin. Microbiol. 5:240245.
98. Tringe, S. G.,, C. von Mering,, A. Kobayashi,, A. A. Salamov,, K. Chen,, H. W. Chang,, M. Podar,, J. M. Short,, E. J. Mathur,, J. C. Detter,, P. Bork,, P. Hugenholtz, and, E. M. Rubin. 2005. Comparative metagenomics of microbial communities. Science 308:554557.
99. Tyson, J. W.,, J. Chapman,, P. Hugenholtz,, V. V. Solovyev,, E. M. Rubin,, D. S. Rokhsar, and, J. F. Banfield. 2004. Community structure and metabolism through reconstruction of microbial genomes from the environment. Nature 428:37-43.
100. Uchiyama, T.,, T. Abe,, T. Ikemura, and, K. Watanabe. 2005. Substrate-induced gene-expression screening of environmental metagenome libraries for isolation of catabolic genes. Nat. Biotechnol. 23:8893.
101. Urbach, E.,, K. L. Vergin, and, S. J. Giovannoni. 1999. Immunochemical detection and isolation of DNA from metabolically active bacteria. Appl. Environ. Microbiol. 65:12071213.
102. Ussery, D. W.,, and P. F. Hallin. 2004. Genome update: length distributions of sequenced prokaryotic genomes. Microbiology 150:513516.
103. Vallaeys, T.,, F. Persello-Cartieaux,, N. Rouard,, C. Lors,, G. Laguerre, and, G. Soulas. 1997. PCR-RFLP analysis of 16S rRNA, tfd A and tfd B genes reveals a diversity of 2,4-D degraders in soil aggregates. FEMS Microbiol. Lett. 24:269278.
104. Venter, J. C.,, K. Remington,, J. F. Heidelberg,, A. L. Halpern,, D. Rush,, J. A. Eisen,, D. Wu,, I. Paulsen,, K. E. Nelson,, W. Nelson,, D. E. Fouts,, S. Sevy,, H. Knap,, M. W. Lomas,, K. Nealson,, O. White,, J. D. Peterson,, J. Hoffman,, R. Parsons,, H. Baden-Tillson,, C. Pfannkoch,, Y. Rogers, and, H. O. Smith. 2004. Environmental genome shotgun sequencing of the Sargasso Sea. Science 304:6674.
105. Watanabe, K.,, M. Teramoto,, H. Futamata, and, S. Harayama. 1998. Molecular detection, isolation, and physiological characterization of functionally dominant phenol-degrading bacteria in activated sludge. Appl. Environ. Microbiol. 64:43964402.
106. Wawrik, B. L.,, G. Kerkhof,, J. Zylstra, and, J. L. Kukor. 2005. Identification of unique type II polyketide synthase genes in soil. Appl. Environ. Microbiol. 71:22322238.
107. Wellington, E. M. H.,, A. Barry, and, M. Krsek. 2003. Resolving functional diversity in relation to microbial community structure in soil; exploiting genomics and stable isotope probing. Curr. Opin. Microbiol. 6:295301.
108. Wen, C. M.,, C. S. Tseng,, C. Y. Cheng, and, Y. K. Li. 2002. Purification, characterization and cloning of a chitinase from Bacillus sp. NCTU2. Biotechnol. Appl. Biochem. 35:213219.
109. Wilson, M. S.,, C. Bakermans, and, E. Madsen. 1999. In situ, real-time catabolic gene expression: extraction and characterization of naphthalene dioxygenase mRNA transcripts from groundwater. Appl. Environ. Microbiol. 65:8087.
110. Wommack, K. E.,, S. R. Bench,, K. E. Williamson, and, M. Radosevich. 2004. Viruses in soils: the first terrestrial viral metagenome, p. 11. In Proceedings of the 10th International Symposium on Microbial Ecology, Cancun, Mexico, August, 2004. Kenes International, Geneva, Switzerland.
111. Wu, Y.,, V. M. Hayes,, J. Osinga,, I. M. Mulder,, M. W. Looman,, C. H. Buys, and, R. M. Hofstra. 1998. Improvement of fragment and primer selection for mutation detection by denaturing gradient gel electrophoresis. Nucleic Acids Res. 26:54325440.
112. Zengler, K.,, G. Toledo,, M. Rappe,, J. Elkins,, E. J. Mathur,, J. M. Short, and, M. Keller. 2002. Cultivating the uncultured. Proc. Natl. Acad. Sci. USA 99:1568115686.
113. Zhou, J.-Z. 2003. Microarrays for bacterial detection and microbial community analysis. Curr. Opin. Microbiol. 6:294.
114. Zhou, J.-Z.,, and D. K. Thompson. 2002. Challenges in applying microarrays to environmental studies. Curr. Opin. Biotechnol. 13:202204.


Generic image for table

Classification of microarrays used in environmental studies

Citation: Cowan D, Stafford W. 2007. Metagenomic Methods for the Identification of Active Microorganisms and Genes in Biodegradation Processes, p 1079-1088. In Hurst C, Crawford R, Garland J, Lipson D, Mills A, Stetzenbach L (ed), Manual of Environmental Microbiology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555815882.ch86

This is a required field
Please enter a valid email address
Please check the format of the address you have entered.
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error