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Nat Protoc. 2008;3(8):1261-9. doi: 10.1038/nprot.2008.102.

Cultivating previously uncultured soil bacteria using a soil substrate membrane system.

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  • 1School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney, New South Wales 2052, Australia.


Most bacteria are recalcitrant to traditional cultivation in the laboratory. The soil substrate membrane system provides a simulated environment for the cultivation of previously undescribed soil bacteria as microcolonies. The system uses a polycarbonate membrane as a solid support for growth and soil extract as the substrate. Diverse microcolonies can be visualized using total bacterial staining combined with fluorescence in situ hybridization (FISH) after 7-10-d incubation. Molecular typing shows that the majority of microcolony-forming bacteria recovered using this protocol were resistant to growth using standard methods. The protocol takes <4 h of bench time over the 10-d period.

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