Using phage display selected antibodies to dissect microbiomes for complete de novo genome sequencing of low abundance microbes

BMC Microbiology

Methodology Article

Cliff S Han¹  Ashlynn R Daughton¹  Devin W Close¹  Timothy C Sanchez¹  Armand EK Dichosa¹  Nileena Velappan¹  Andrew RM Bradbury¹  Csaba Kiss¹  Hajnalka E Daligault¹  Krista G Reitenga¹  Srinivas Iyer¹  Sandeep Kumar¹  Fortunato Ferrara² 

[1] Bioscience Division, Los Alamos National Laboratory, Los Alamos, NM, USA;New Mexico Consortium, Los Alamos, NM, USA;

关键词: Phage antibodies;    Genome completion;    Single cell genomics;    MDA;    Flow cytometry;   

DOI  :  10.1186/1471-2180-13-270

 received in 2013-08-31, accepted in 2013-11-21,  发布年份 2013

来源: Springer

【 摘 要 】

BackgroundSingle cell genomics has revolutionized microbial sequencing, but complete coverage of genomes in complex microbiomes is imperfect due to enormous variation in organismal abundance and amplification bias. Empirical methods that complement rapidly improving bioinformatic tools will improve characterization of microbiomes and facilitate better genome coverage for low abundance microbes.MethodsWe describe a new approach to sequencing individual species from microbiomes that combines antibody phage display against intact bacteria with fluorescence activated cell sorting (FACS). Single chain (scFv) antibodies are selected using phage display against a bacteria or microbial community, resulting in species-specific antibodies that can be used in FACS for relative quantification of an organism in a community, as well as enrichment or depletion prior to genome sequencing.ResultsWe selected antibodies against Lactobacillus acidophilus and demonstrate a FACS-based approach for identification and enrichment of the organism from both laboratory-cultured and commercially derived bacterial mixtures. The ability to selectively enrich for L. acidophilus when it is present at a very low abundance (<0.2%) leads to complete (>99.8%) de novo genome coverage whereas the standard single-cell sequencing approach is incomplete (<68%). We show that specific antibodies can be selected against L. acidophilus when the monoculture is used as antigen as well as when a community of 10 closely related species is used demonstrating that in principal antibodies can be generated against individual organisms within microbial communities.ConclusionsThe approach presented here demonstrates that phage-selected antibodies against bacteria enable identification, enrichment of rare species, and depletion of abundant organisms making it tractable to virtually any microbe or microbial community. Combining antibody specificity with FACS provides a new approach for characterizing and manipulating microbial communities prior to genome sequencing.

【 授权许可】

Unknown   
© Close et al.; licensee BioMed Central Ltd. 2013. This article is published under license to BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.