Please use this identifier to cite or link to this item: https://hdl.handle.net/10356/142441
Title: Primer-free FISH probes from metagenomics/metatranscriptomics data permit the study of uncharacterised taxa in complex microbial communities
Authors: Tan, Shi Ming
Yung, Maria Pui Yi
Hutchinson, Paul E.
Xie, Chao
Teo, Guo Hui
Muhammad Hafiz Ismail
Drautz-Moses, Daniela Isabel
Little, Peter F. R.
Williams, Rohan B. H.
Cohen, Yehuda
Keywords: Science::Biological sciences::Microbiology
Issue Date: 2019
Source: Tan, S. M., Yung, M. P. Y., Hutchinson, P. E., Xie, C., Teo, G. H., Muhammad Hafiz Ismail, . . . Cohen, Y. (2019). Primer-free FISH probes from metagenomics/metatranscriptomics data permit the study of uncharacterised taxa in complex microbial communities. npj Biofilms and Microbiomes, 5(1), 17-. doi:10.1038/s41522-019-0090-9
Journal: npj Biofilms and Microbiomes
Abstract: Methods for the study of member species in complex microbial communities remain a high priority, particularly for rare and/or novel member species that might play an important ecological role. Specifically, methods that link genomic information of member species with its spatial structure are lacking. This study adopts an integrative workflow that permits the characterisation of previously unclassified bacterial taxa from microbiomes through: (1) imaging of the spatial structure; (2) taxonomic classification and (3) genome recovery. Our study attempts to bridge the gaps between metagenomics/metatranscriptomics and high-resolution biomass imaging methods by developing new fluorescence in situ hybridisation (FISH) probes—termed as R-Probes—from shotgun reads that harbour hypervariable regions of the 16S rRNA gene. The sample-centric design of R-Probes means that probes can directly hybridise to OTUs as detected in shotgun sequencing surveys. The primer-free probe design captures larger microbial diversity as compared to canonical probes. R-Probes were designed from deep-sequenced RNA-Seq datasets for both FISH imaging and FISH–Fluorescence activated cell sorting (FISH–FACS). FISH–FACS was used for target enrichment of previously unclassified bacterial taxa prior to downstream multiple displacement amplification (MDA), genomic sequencing and genome recovery. After validation of the workflow on an axenic isolate of Thauera species, the techniques were applied to investigate two previously uncharacterised taxa from a tropical full-scale activated sludge community. In some instances, probe design on the hypervariable region allowed differentiation to the species level. Collectively, the workflow can be readily applied to microbiomes for which shotgun nucleic acid survey data is available.
URI: https://hdl.handle.net/10356/142441
ISSN: 2055-5008
DOI: 10.1038/s41522-019-0090-9
Rights: © 2019 The Author(s). Published in partnership with Nanyang Technological University. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
Fulltext Permission: open
Fulltext Availability: With Fulltext
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