Benchmarking of single-virus genomics: a new tool for uncovering the virosphere
Inmaculada Garcia-Heredia
Department of Physiology, Genetics, and Microbiology, University of Alicante, Alicante, Spain
Search for more papers by this authorAnanda S. Bhattacharjee
Bigelow Laboratory for Ocean Sciences, East Boothbay, ME, USA
Search for more papers by this authorOscar Fornas
Flow Cytometry Unit, Centre for Genomic Regulation (CRG), The Barcelona Institute for Science and Technology (BIST), Barcelona, Spain
Flow Cytometry Unit, Departament de Ciències Experimentals i de la Salut, Universitat Pompeu Fabra (UPF), Barcelona, Spain
Search for more papers by this authorMonica L. Gomez
Department of Physiology, Genetics, and Microbiology, University of Alicante, Alicante, Spain
Search for more papers by this authorJoaquín M. Martínez
Bigelow Laboratory for Ocean Sciences, East Boothbay, ME, USA
Search for more papers by this authorCorresponding Author
Manuel Martinez-Garcia
Department of Physiology, Genetics, and Microbiology, University of Alicante, Alicante, Spain
For correspondence. E-mail [email protected], Tel: (+34) 9653853; Fax: (+34) 96 590 9569.
Search for more papers by this authorInmaculada Garcia-Heredia
Department of Physiology, Genetics, and Microbiology, University of Alicante, Alicante, Spain
Search for more papers by this authorAnanda S. Bhattacharjee
Bigelow Laboratory for Ocean Sciences, East Boothbay, ME, USA
Search for more papers by this authorOscar Fornas
Flow Cytometry Unit, Centre for Genomic Regulation (CRG), The Barcelona Institute for Science and Technology (BIST), Barcelona, Spain
Flow Cytometry Unit, Departament de Ciències Experimentals i de la Salut, Universitat Pompeu Fabra (UPF), Barcelona, Spain
Search for more papers by this authorMonica L. Gomez
Department of Physiology, Genetics, and Microbiology, University of Alicante, Alicante, Spain
Search for more papers by this authorJoaquín M. Martínez
Bigelow Laboratory for Ocean Sciences, East Boothbay, ME, USA
Search for more papers by this authorCorresponding Author
Manuel Martinez-Garcia
Department of Physiology, Genetics, and Microbiology, University of Alicante, Alicante, Spain
For correspondence. E-mail [email protected], Tel: (+34) 9653853; Fax: (+34) 96 590 9569.
Search for more papers by this authorSummary
Metagenomics and single-cell genomics have enabled the discovery of relevant uncultured microbes. Recently, single-virus genomics (SVG), although still in an incipient stage, has opened new avenues in viral ecology by allowing the sequencing of one single virus at a time. The investigation of methodological alternatives and optimization of existing procedures for SVG is paramount to deliver high-quality genomic data. We report a sequencing dataset of viral single-amplified genomes (vSAGs) from cultured and uncultured viruses obtained by applying different conditions in each SVG step, from viral preservation and novel whole-genome amplification (WGA) to sequencing platforms and genome assembly. Sequencing data showed that cryopreservation and mild fixation were compatible with WGA, although fresh samples delivered better genome quality data. The novel TruPrime WGA, based on primase-polymerase features, and WGA-X employing a thermostable phi29 polymerase, were proven to be with sufficient sensitivity in SVG. The Oxford Nanopore (ON) sequencing platform did not provide a significant improvement of vSAG assembly compared to Illumina alone. Finally, the SPAdes assembler performed the best. Overall, our results represent a valuable genomic dataset that will help to standardized and advance new tools in viral ecology.
Supporting Information
| Filename | Description |
|---|---|
| emi15375-sup-0001-supinfo.xlsxExcel 2007 spreadsheet , 27.1 KB | Data S1. Supporting information. |
| emi15375-sup-0002-Figures.docxWord 2007 document , 123.3 KB | Fig. S1. Comparison of two whole-genome amplification methods for single cells of E. coli BL21. A. Boxplot comparing the means of the bases covered of E. coli BL21 genome/MB sequenced of each SC. B. Dot plot of bases covered of E. coli BL21 genome/MB sequenced for each SC. C. Box plot comparing the means of length (bp) of the longest contigs obtained for each assembler. Assembly was performed applying same parameters as for single viruses. Fig. S2. Comparison of preservation methods for sequenced bacteriophagesT4vSAG. Box plot comparing the average coverage obtained for each preservation method. |
| emi15375-sup-0003-Tables.docxWord 2007 document , 19.2 KB | Table S1. ANOVA and Tukey's test comparing conservation methods. Table S2. ANOVA and Tukey's test comparing assembly programs. |
Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.
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