Plant Soil Environ., 2009, 55(10):413-423 | DOI: 10.17221/132/2009-PSE

Denaturing gradient gel electrophoresis as a fingerprinting method for the analysis of soil microbial communities

V. Valášková, P. Baldrian
Laboratory of Environmental Microbiology, Institute of Microbiology of the Academy of Sciences of the Czech Republic, Prague, Czech Republic

In soil microbial ecology, the effects of environmental factors and their gradients, temporal changes or the response to specific experimental treatments of microbial communities can only be effectively analyzed using methods that address the structural differences among whole communities. Fingerprinting methods are the most appropriate technique for this task when multiple samples must be analyzed. Among the methods currently used to compare microbial communities based on nucleic acid sequences, the techniques based on differences in the melting properties of double-stranded molecules, denaturing gradient gel electrophoresis (DGGE) or temperature gradient gel electrophoresis (TGGE), are the most widely used. Their main advantage is that they provide the possibility to further analyze whole sequences contained in fingerprints using molecular methods. In addition to the analysis of microbial communities based on DNA extracted from soils, DGGE/TGGE can also be used for the assessment of the active part of the community based on the analysis of RNA-derived sequences or for the analysis of sequences of functional genes encoding for proteins involved in important soil processes.

Keywords: DGGE; bacteria; fungi; microbial community; soil ecology; TGGE

Published: October 31, 2009  Show citation

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Valášková V, Baldrian P. Denaturing gradient gel electrophoresis as a fingerprinting method for the analysis of soil microbial communities. Plant Soil Environ. 2009;55(10):413-423. doi: 10.17221/132/2009-PSE.
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    References

    1. Anderson I.C., Cairney J.W.G. (2004): Diversity and ecology of soil fungal communities: increased understanding through the application of molecular techniques. Environmental Microbiology, 6: 769-779. Go to original source... Go to PubMed...
    2. Anderson I.C., Campbell C.D., Prosser J.I. (2003): Diversity of fungi in organic soils under a moorland - Scots pine (Pinus sylvestris L.) gradient. Environmental Microbiology, 5: 1121-1132. Go to original source... Go to PubMed...
    3. Aneja M.K., Sharma S., Munch J.C., Schloter M. (2004): RNA fingerprinting - a new method to screen for differences in plant litter degrading microbial communities. Journal of Microbiological Methods, 59: 223-231. Go to original source... Go to PubMed...
    4. Arenz B.E., Held B.W., Jurgens J.A., Farrell R.L., Blanchette R.A. (2006): Fungal diversity in soils and historic wood from the Ross Sea Region of Antarctica. Soil Biology and Biochemistry, 38: 3057-3064. Go to original source...
    5. Artz R.R.E., Anderson I.C., Chapman S.J., Hagn A., Schloter M., Potts J.M., Campbell C.D. (2007): Changes in fungal community composition in response to vegetational succession during the natural regeneration of cutover peatlands. Microbial Ecology, 54: 508-522. Go to original source... Go to PubMed...
    6. Baldrian P., Trögl J., Frouz J., Šnajdr J., Valášková V., Merhautová V., Cajthaml T., Herinková J. (2008): Enzyme activities and microbial biomass in topsoil layer during spontaneous succession in spoil heaps after brown coal mining. Soil Biology and Biochemistry, 40: 2107-2115. Go to original source...
    7. Bastias B.A., Anderson I.C., Xu Z., Cairney J.W.G. (2007): RNA- and DNA-based profiling of soil fungal communities in a native Australian eucalypt forest and adjacent Pinus elliotti plantation. Soil Biology and Biochemistry, 39: 3108-3114. Go to original source...
    8. Bougoure D.S., Cairney J.W.G. (2005): Fungi associated with hair roots of Rhododendron lochiae (Ericaceae) in an Australian tropical cloud forest revealed by culturing and culture-independent molecular methods. Environmental Microbiology, 7: 1743-1754. Go to original source... Go to PubMed...
    9. Clegg C.D. (2006): Impact of cattle grazing and inorganic fertiliser additions to managed grasslands on the microbial community composition of soils. Applied Soil Ecology, 31: 73-82. Go to original source...
    10. Diouf M., Miambi E., Mora P., Delgarde S., Rouland C. (2006): The impact of termite sheetings age on their fungal communities. European Journal of Soil Biology, 42: 85-91. Go to original source...
    11. Dohrmann A.B., Tebbe C.C. (2004): Microbial community analysis by PCR-single-strand conformation polymorphism (PCR-SSCP). In: Kowalchuk G.A., de Bruijn F.J., Head I.M., Akkermans A.D.L., van Elsas J.D. (eds): Molecular Microbial Ecology Manual. Kluwer Academic Publishers, Dordrecht, 809-838.
    12. Duineveld B.M., Kowalchuk G.A., Keijzer A., van Elsas J.D., van Veen J.A. (2001): Analysis of bacterial communities in the rhizosphere of chrysanthemum via denaturing gradient gel electrophoresis of PCR-amplified 16S rRNA as well as DNA fragments coding for 16S rRNA. Applied and Environmental Microbiology, 67: 172-178. Go to original source... Go to PubMed...
    13. Felske A., Engelen B., Nübel U., Backhaus H. (1996): Direct ribosome isolation from soil to extract bacterial rRNA for community analysis. Applied and Environmental Microbiology, 62: 3034-3036. Go to original source... Go to PubMed...
    14. Findlay R.H., King G.M., Watling L. (1989): Efficacy of phospholipid analysis in determining microbial biomass in sediments. Applied and Environmental Microbiology, 55: 2888-2893. Go to original source... Go to PubMed...
    15. Fioretto A., Papa S., Curcio E., Sorrentino G., Fuggi A. (2000): Enzyme dynamics on decomposing leaf litter of Cistus incanus and Myrtus communis in a Mediterranean ecosystem. Soil Biology and Biochemistry, 32: 1847-1855. Go to original source...
    16. Fisher M.M., Triplett E.W. (1999): Automated approach for ribosomal intergenic spacer analysis of microbial diversity and its application to freshwater bacterial communities. Applied and Environmental Microbiology, 65: 4630-4636. Go to original source... Go to PubMed...
    17. Gardes M., Bruns T.D. (1993): ITS primers with enhanced specificity for basidiomycetes - application to the identification of mycorrhizae and rusts. Molecular Ecology, 2: 113-118. Go to original source... Go to PubMed...
    18. Gelsomino A., Cacco G. (2006): Compositional shifts of bacterial groups in a solarized and amended soil as determined by denaturing gradient gel electrophoresis. Soil Biology and Biochemistry, 38: 91-102. Go to original source...
    19. Gremion F., Chatzinotas A., Kaufmann K., Von Sigler W., Harms H. (2004): Impacts of heavy metal contamination and phytoremediation on a microbial community during a twelve-month microcosm experiment. FEMS Microbiology Ecology, 48: 273-283. Go to original source... Go to PubMed...
    20. Griffiths R.I., Manefield M., Whiteley A.S., Bailey M.J. (2004): DNA and RNA extraction from soil. In: Kowalchuk G.A., de Bruijn F.J., Head I.M., Akkermans A.D.L., van Elsas J.D. (eds): Molecular Microbial Ecology Manual. Kluwer Academic Publishers, Dordrecht, 149-158.
    21. Hernesmaa A., Björklöf K., Kiikkilä O., Fritze H., Haahtela K., Romantschuk M. (2005): Structure and function of microbial communities in the rhizosphere of Scots pine after tree-felling. Soil Biology and Biochemistry, 37: 777-785. Go to original source...
    22. Heuer H., Smala K. (1997): Application of denaturating gradient gel electrophoresis (DGGE) and temperature gradient gel epectrophoresis (TGGE) for studying soil microbial communities. In: van Elsas J.D., Wellington E.M.H., Trevors J.T. (eds): Modern Soil Microbiology. Marcel Dekker New York, 353-373.
    23. Heuer H., Krsek M., Baker P., Smalla K., Wellington E.M.H. (1997): Analysis of actinomycete communities by specific amplification of genes encoding 16S rRNA and gel-electrophoretic separation in denaturing gradients. Applied and Environmental Microbiology, 63: 3233-3241. Go to original source... Go to PubMed...
    24. Jaatinen K., Laiho R., Vuorenmaa A., del Castillo U., Minkkinen K., Pennanen T., Penttila T., Fritze H. (2008): Responses of aerobic microbial communities and soil respiration to water-level drawdown in a northern boreal fen. Environmental Microbiology, 10: 339-353. Go to original source... Go to PubMed...
    25. Kandeler E., Tscherko D., Spiegel H. (1999): Long-term monitoring of microbial biomass, N mineralization and enzyme activities of a Chernozem under different tillage management. Biology and Fertility of Soils, 28: 343-351. Go to original source...
    26. Korkama-Rajala T., Mueller M.M., Pennanen T. (2008): Decomposition and fungi of needle litter from slowand fast-growing norway spruce (Picea abies) clones. Microbial Ecology, 56: 76-89. Go to original source... Go to PubMed...
    27. Kowalchuk G.A., Smit E. (2004): Fungal community analysis using PCR-denaturing gradient gel electrophoresis (DGGE). In: Kowalchuk G.A., de Bruijn F.J., Head I.M., Akkermans A.D.L., van Elsas J.D. (eds): Molecular Microbial Ecology Manual. Kluwer Academic Publishers, Dordrecht, 771-788.
    28. Kowalchuk G.A., Gerards S., Woldendorp J.W. (1997): Detection and characterization of fungal infections of Ammophila arenaria (marram grass) roots by denaturing gradient gel electrophoresis of specifically amplified 18S rDNA. Applied and Environmental Microbiology, 63: 3858-3865. Go to original source... Go to PubMed...
    29. Kowalchuk G.A., de Souza F.A., van Veen J.A. (2002): Community analysis of arbuscular mycorrhizal fungi associated with Ammophila arenaria in Dutch coastal sand dunes. Molecular Ecology, 11: 571-581. Go to original source... Go to PubMed...
    30. Kowalchuk G.A., Bodelier P.L.E., Heilig G.H.J., Stephen J.R., Laanbroek H.J. (1998): Community analysis of ammonia-oxidising bacteria, in relation to oxygen availability in soils and root-oxygenated sediments, using PCR, DGGE and oligonucleotide probe hybridisation. FEMS Microbiology Ecology, 27: 339-350. Go to original source...
    31. Kowalchuk G.A., de Bruijn F.J., Head I.M., Akkermans A.D., van Elsas J.D. (eds.) (2004): Molecular Microbial Ecology Manual. Kluwer Academic Publishers, Dordrecht. Go to original source...
    32. Kozdrój J., van Elsas J.D. (2001): Structural diversity of microbial communities in arable soils of a heavily industrialised area determined by PCR-DGGE fingerprinting and FAME profiling. Applied Soil Ecology, 17: 31-42. Go to original source...
    33. Larena I., Salazar O., Gonzalez V., Julian M.C., Rubio V. (1999): Design of a primer for ribosomal DNA internal transcribed spacer with enhanced specificity for ascomycetes. Journal of Biotechnology, 75: 187-194. Go to original source... Go to PubMed...
    34. Lee D.H., Zo Y.G., Kim S.J. (1996): Nonradioactive method to study genetic profiles of natural bacterial communities by PCR-single-strand-conformation polymorphism. Applied and Environmental Microbiology, 62: 3112-3120. Go to original source... Go to PubMed...
    35. Liu W.T., Marsh T.L., Cheng H., Forney L.J. (1997): Characterization of microbial diversity by determining terminal restriction fragment length polymorphisms of genes encoding 16S rRNA. Applied and Environmental Microbiology, 63: 4516-4522. Go to original source... Go to PubMed...
    36. Marschner P., Kandeler E., Marschner B. (2003): Structure and function of the soil microbial community in a long-term fertilizer experiment. Soil Biology and Biochemistry, 35: 453-461. Go to original source...
    37. May L.A., Smiley B., Schmidt M.G. (2001): Comparative denaturing gradient gel electrophoresis analysis of fungal communities associated with whole plant corn silage. Canadian Journal of Microbiology, 47: 829-841. Go to original source... Go to PubMed...
    38. Möhlenhoff P., Muller L., Gorbushina A.A., Petersen K. (2001): Molecular approach to the characterisation of fungal communities: methods for DNA extraction, PCR amplification and DGGE analysis of painted art objects. FEMS Microbiology Letters, 195: 169-173. Go to original source... Go to PubMed...
    39. Muyzer G., de Waal E.C., Uitterlinden A.G. (1993): Profiling of complex microbial populations by denaturing gradient gel electrophoresis analysis of polymerase chain reaction-amplified genes coding for 16S ribosomal RNA. Applied and Environmental Microbiology, 59: 695-700. Go to original source... Go to PubMed...
    40. Muyzer G., Teske A., Wirsen C.O., Jannasch H.W. (1995): Phylogenetic relationships of Thiomicrospira species and their identification in deep-sea hydrothermal vent samples by denaturing gradient gel electrophoresis of 16S rDNA fragments. Archives of Microbiology, 164: 165-171. Go to original source... Go to PubMed...
    41. Muyzer G., Brinkhoff T., Nubel U., Santegoeds C., Schafer H., Wawer C. (2004): Denaturing gradient gel electrophoresis (DGGE) in microbial ecology. In: Kowalchuk G.A., de Bruijn F.J., Head I.M., Akkermans A.D.L., van Elsas J.D. (eds): Molecular Microbial Ecology Manual. Kluwer Academic Publishers Dordrecht, 743-769.
    42. Oros-Sichler M., Gomes N.C.M., Neuber G., Smalla K. (2006): A new semi-nested PCR protocol to amplify large 18S rRNA gene fragments for PCR-DGGE analysis of soil fungal communities. Journal of Microbiological Methods, 65: 63-75. Go to original source... Go to PubMed...
    43. Oros-Sichler M., Costa R., Heuer H., Smalla K. (2007): Molecular Fingerprinting Techniques to Analyze Soil Microbial Communities. In: van Elsas J.D., Jansson J.K., Trevors J.T. (eds): Modern Soil Microbiology. CRC Press Madison, 355-386.
    44. Pennanen T., Caul S., Daniell T.J., Griffiths B.S., Ritz K., Wheatley R.E. (2004): Community-level responses of metabolically-active soil microorganisms to the quantity and quality of substrate inputs. Soil Biology and Biochemistry, 36: 841-848. Go to original source...
    45. Phillips L.A., Germida J.J., Farrell R.E., Greer C.W. (2008): Hydrocarbon degradation potential and activity of endophytic bacteria associated with prairie plants. Soil Biology and Biochemistry, 40: 3054-3064. Go to original source...
    46. Riesner D., Henco K., Steger G. (1990): Temperature-Gradient Gel Electrophoresis: A method for the analysis of conformational transitions and mutations in nucleic acids and protein. In: Chrambach A., Dunn M.J., Radola B.J. (eds): Advances in Electrophoresis, Vol. 4. VCH Verlagsgesellschaft, Weinheim, 169-250.
    47. Roesch L.F., Fulthorpe R.R., Riva A., Casella G., Hadwin A.K.M., Kent A.D., Daroub S.H., Camargo F.A.O., Farmerie W.G., Triplett E.W. (2007): Pyrosequencing enumerates and contrasts soil microbial diversity. International Society for Microbial Ecology Journal, 1: 283-290. Go to original source... Go to PubMed...
    48. Ros M., Pascual J.A., Garcia C., Hernandez M.T., Insam H. (2006): Hydrolase activities, microbial biomass and bacterial community in a soil after long-term amendment with different composts. Soil Biology and Biochemistry, 38: 3443-3452. Go to original source...
    49. Rosset R., Julien J., Monier R. (1966): Ribonucleic acid composition of bacteria as a function of growth rate. Journal of Molecular Biology, 18: 308-320. Go to original source... Go to PubMed...
    50. Saetre P., Baath E. (2000): Spatial variation and patterns of soil microbial community structure in a mixed spruce-birch stand. Soil Biology and Biochemistry, 32: 909-917. Go to original source...
    51. Sagova-Mareckova M., Cermak L., Novotna J., Plhackova K., Forstova J., Kopecky J. (2008): Innovative methods for soil DNA purification tested in soils with widely differing characteristics. Applied and Environmental Microbiology, 74: 2902-2907. Go to original source... Go to PubMed...
    52. Sakurai M., Suzuki K., Onodera M., Shinano T., Osaki M. (2007): Analysis of bacterial communities in soil by PCR-DGGE targeting protease genes. Soil Biology and Biochemistry, 39: 2777-2784. Go to original source...
    53. Santos J.C., Finlay R.D., Tehler A. (2006): Molecular analysis of arbuscular mycorrhizal fungi colonising a semi-natural grassland along a fertilisation gradient. New Phytologist, 172: 159-168. Go to original source... Go to PubMed...
    54. Seifert K.A., Samson R.A., Dewaard J.R., Houbraken J., Levesque C.A., Moncalvo J.M., Louis-Seize G., Hebert P.D.N. (2007): Prospects for fungus identification using C01 DNA barcodes, with Penicillium as a test case. Proceedings of the National Academy of Sciences of the United States of America, 104: 3901-3906. Go to original source... Go to PubMed...
    55. Shalon D., Smith S.J., Brown P.O. (1996): A DNA microarray system for analyzing complex DNA samples using two-color fluorescent probe hybridization. Genome Research, 6: 639-645. Go to original source... Go to PubMed...
    56. Smit E., Leeflang P., Glandorf B., van Elsas J.D., Wernars K. (1999): Analysis of fungal diversity in the wheat rhizosphere by sequencing of cloned PCR-amplified genes encoding 18S rRNA and temperature gradient gel electrophoresis. Applied and Environmental Microbiology, 65: 2614-2621. Go to original source... Go to PubMed...
    57. Šnajdr J., Valášková V., Merhautová V., Herinková J., Cajthaml T., Baldrian P. (2008): Spatial variability of enzyme activities and microbial biomass in the upper layers of Quercus petraea forest soil. Soil Biology and Biochemistry, 40: 2068-2075. Go to original source...
    58. Vainio E.J., Hantula J. (2000): Direct analysis of woodinhabiting fungi using denaturing gradient gel electrophoresis of amplified ribosomal DNA. Mycological Research, 104: 927-936. Go to original source...
    59. Valášková V., Šnajdr J., Bittner B., Cajthaml T., Merhautová V., Hofrichter M., Baldrian P. (2007): Production of lignocellulose-degrading enzymes and degradation of leaf litter by saprotrophic basidiomycetes isolated from a Quercus petraea forest. Soil Biology and Biochemistry, 39: 2651-2660. Go to original source...
    60. van Elsas J.D., Duarte G.F., Keijzer-Wolters A., Smit E. (2000): Analysis of the dynamics of fungal communities in soil via fungal-specific PCR of soil DNA followed by denaturing gradient gel electrophoresis. Journal of Microbiological Methods, 43: 133-151. Go to original source... Go to PubMed...
    61. Wartiainen I., Eriksson T., Zheng W., Rasmussen U. (2008): Variation in the active diazotrophic community in rice paddy-nifH PCR-DGGE analysis of rhizosphere and bulk soil. Applied Soil Ecology, 39: 65-75. Go to original source...
    62. White T.J., Bruns T., Lee S., Taylor J. (1990): Amplification and direct sequencing of fungal ribosomal RNA genes for phylogenetics. In: Innins M.A., Gelfand D.H., Sninsky J.J., White T.J. (eds): PCR Protocols. Academic Press San Diego, 315-322. Go to original source...
    63. Yao S., Merwin I.A., Abawi G.S., Thies J.E. (2006): Soil fumigation and compost amendment alter soil microbial community composition but do not improve tree growth or yield in an apple replant site. Soil Biology and Biochemistry, 38: 587-599. Go to original source...
    64. Zhou J.Z., Bruns M.A., Tiedje J.M. (1996): DNA recovery from soils of diverse composition. Applied and Environmental Microbiology, 62: 316-322. Go to original source... Go to PubMed...
    65. Zuccaro A., Schulz B., Mitchell J.I. (2003): Molecular detection of ascomycetes associated with Fucus serratus. Mycological Research, 107: 1451-1466. Go to original source... Go to PubMed...

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