Publikationsdatum:
2022-05-25
Beschreibung:
© The Author(s), 2013. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in PLoS ONE 8 (2013): e76096, doi:10.1371/journal.pone.0076096.
Beschreibung:
DNA samples derived from vertebrate skin, bodily cavities and body fluids contain both host and microbial DNA; the latter often present as a minor component. Consequently, DNA sequencing of a microbiome sample frequently yields reads originating from the microbe(s) of interest, but with a vast excess of host genome-derived reads. In this study, we used a methyl-CpG binding domain (MBD) to separate methylated host DNA from microbial DNA based on differences in CpG methylation density. MBD fused to the Fc region of a human antibody (MBD-Fc) binds strongly to protein A paramagnetic beads, forming an effective one-step enrichment complex that was used to remove human or fish host DNA from bacterial and protistan DNA for subsequent sequencing and analysis. We report enrichment of DNA samples from human saliva, human blood, a mock malaria-infected blood sample and a black molly fish. When reads were mapped to reference genomes, sequence reads aligning to host genomes decreased 50-fold, while bacterial and Plasmodium DNA sequences reads increased 8–11.5-fold. The Shannon-Wiener diversity index was calculated for 149 bacterial species in saliva before and after enrichment. Unenriched saliva had an index of 4.72, while the enriched sample had an index of 4.80. The similarity of these indices demonstrates that bacterial species diversity and relative phylotype abundance remain conserved in enriched samples. Enrichment using the MBD-Fc method holds promise for targeted microbiome sequence analysis across a broad range of sample types.
Beschreibung:
LAZ and VS were funded by a Brown University Office of the Vice President of Research SEED grant (LAZ) entitled “Tracking disease spread through the wildlife trade: New techniques to identify infectious microbes in aquarium fishes.” SOO and MQ were funded by Wellcome Trust Sanger Institute grant numbers 098051 and 079355/Z/06/Z.
Repository-Name:
Woods Hole Open Access Server
Materialart:
Article
Format:
image/tiff
Format:
application/pdf
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text/csv
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