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Authors: Bonnie Poulos
Group: VERVE Net, Sullivan Lab

Proper citation: Bonnie Poulos 2016. Pseudoalteromonas Media Recipes. protocols.io https://dx.doi.org/10.17504/protocols.io.ek6bcze Copy   

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Authors: Seth John, Bonnie Poulos, Christine Schirmer
Group: VERVE Net, Sullivan Lab
Summary: This protocol describes a technique to recover viruses from natural waters using iron- based flocculation and large-pore-size filtration, followed by resuspension of virus- containing precipitates in a pH 6.5 buffer. This Fe-based virus flocculation, filtration and resuspension method (FFR) is efficient (> 90% recovery), reliable, inexpensive and adaptable to many aspects of marine viral ecology and genomics research. Recovered viruses are amenable to gene sequencing, and a variable proportion of phages, depending upon the phage, retain their infectivity when recovered if using oxalic acid in the resuspension buffer. Particles lose infectivity if resuspended with ascorbic acid in the buffer.

Proper citation: Seth John, Bonnie Poulos, Christine Schirmer 2015. Iron Chloride Precipitation of Viruses from Seawater. protocols.io https://dx.doi.org/10.17504/protocols.io.c2wyfd Copy   

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Authors: Li Deng
Group: VERVE Net, Sullivan Lab
Summary: Formulation for a phosphate-buffered 20% glutaraldehyde stock solution.

Proper citation: Li Deng 2015. Buffered glutaraldehyde. protocols.io https://dx.doi.org/10.17504/protocols.io.c7gzjv Copy   

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Authors: Matthew Sullivan
Group: VERVE Net, Sullivan Lab

Proper citation: Matthew Sullivan 2016. Calculating burst size. protocols.io https://dx.doi.org/10.17504/protocols.io.ddm245 Copy   

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Authors: Kristin Corrier/Nathan VerBerkmoes
Group: VERVE Net, Sullivan Lab
Summary: For use in "FASP Kit Protocol-ORNL Developed for Bacteriophage"

Proper citation: Kristin Corrier/Nathan VerBerkmoes 2016. Iodoacetamide Solution. protocols.io https://dx.doi.org/10.17504/protocols.io.dej3cm Copy   

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Authors: John H. Paul and Markus Weinbauer
Group: VERVE Net
Summary: This protocol gives a method for prophage induction in marine Synechococcus.Paul, J. H., and M. Weinbauer. 2010. Detection of lysogeny in marine environments, p. 30–33. In S. W. Wilhelm, M. G. Weinbauer, and C. A. Suttle [eds.], Manual of Aquatic Viral Ecology. ASLO.

Proper citation: John H. Paul and Markus Weinbauer 2016. Prophage induction in marine Synechococcus. protocols.io https://dx.doi.org/10.17504/protocols.io.dpc5iv Copy   

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Authors: Matthew Sullivan
Group: VERVE Net, Sullivan Lab

Proper citation: Matthew Sullivan 2016. DNA Extraction Protocol. protocols.io https://dx.doi.org/10.17504/protocols.io.c32yqd Copy   

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Authors: Mathias Middelboe, Amy M. Chan, and Sif K. Bertelsen
Group: VERVE Net, Suttle Laboratory of Marine Molecular Microbiology and Virology
Summary: A generally more efficient way of isolating lytic phages from marine environments is by the use of enrichment cultures. In this approach, the prefiltered water sample that is to be screened for phages against a given target bacterium, is enriched with a bacterial growth medium and amended with that target bacteria (Eisenstark 1967; Carlson 2005). This allows any lytic phages present in the sample to infect the target bacteria and propagate in the cultures, and subsequently, be isolated and purified. The two main advantages of the enrichment approach are 1) that it allows for screening for phages in a much larger volume of sample (typically 25–50 mL, rather than 5–10 µL), thus increasing the probability of isolating rare phages, and 2) that it allows the combination of different target hosts (e.g., different strains of a specific bacteria of interest) in the same incubation, again increasing the possibility of phage isolation.

Proper citation: Mathias Middelboe, Amy M. Chan, and Sif K. Bertelsen 2016. Bacteriophage isolation using enrichment cultures. protocols.io https://dx.doi.org/10.17504/protocols.io.dpt5nm Copy   

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Authors: HANNIGAN GD, GRICE EA, ET AL.
Group: VERVE Net, Club Grice
Summary: This protocol provides method for KEGG Analysis, Gene Ontology Analysis, and Operational Protein Family Analysis. Based on the methods from the following publication: Hannigan, Geoffrey D., et al. "The Human Skin Double-Stranded DNA Virome: Topographical and Temporal Diversity, Genetic Enrichment, and Dynamic Associations with the Host Microbiome." mBio 6.5 (2015): e01578-15.

Proper citation: HANNIGAN GD, GRICE EA, ET AL. 2016. Script P11: Functional Analysis. protocols.io https://dx.doi.org/10.17504/protocols.io.ehubb6w Copy   

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Authors: James E. Thornton
Group: VERVE Net, Sullivan Lab, Culley Lab_ULaval
Summary: This protocol describes the preparation of and treatment with DNase I to degrade DNA in solutions containing iron chloride, EDTA–Mg ascorbate buffer, and cesium chloride. DNase I treatment, CsCl purification, and sucrose purification methods were compared using replicated viral metagenomics in Hurwitz et al. 2012. We use this protocol primarily to remove free host and other contaminating DNA from phage preparations prior to extracting nucleic acid from the phage particles.

Proper citation: James E. Thornton 2015. DNase I Treatment. protocols.io https://dx.doi.org/10.17504/protocols.io.c3myk5 Copy   

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Authors: Ana Del Arco, Noemi Woltermann, Lutz Becks
Group: VERVE Net, Protist Research to Optimize Tools in Genetics (PROT-G)
Summary: Chemostats and other flow through culture systems are a powerful tool for the study of microbial and plankton communities in experimental ecology and evolutionary studies. Commercially available chemostat systems allow the control of a large number of parameters (e.g., ph, pressure, CO2 concentration) but are often expensive and offer a high level of control that is often not needed for many experimental studies. Non-commercial chemostats are more cost efficient, easily set up and flexible in volumes used. Different from semi-continuous culture systems, the flow through conditions of chemostats allow a constant inflow of resources from a reservoir (medium bottle) and outflow of unused nutrients, waste products and organisms, which are all collected in a waste bottle. Nutrient levels in the reservoir and the flow rate of the chemostat system (often referred to as dilution rate and presented as the fraction of the volume of the chemostat that is replaced per day) determines population growth rates and dynamics (for the theory behind chemostats see Smith & Waltman, 1995 and Weitz, 2015). Chemostats have been used in a number of studies and with different organisms and combinations of organisms. For example, Boer et al. (2010) studied how growth-limiting intracellular metabolites control yeast growth under diverse nutrient availability (Saccharomyces cerevisiae growing under five different nutrient supply of nitrogen: carbon: phosphorus). Becks et al. (2005) used chemostat systems to show how changes in the flow rate of a chemostat system influences the population dynamics of a three species microbial system. Frickel et al. (2016) used chemostats to investigate eco- evolutionary dynamics in a coevolving host-virus system (algae Chlorella variabilis and Chlorovirus strain PBCV-1) for 90 days.We present here an instruction for a cost efficient and flexible chemostat systems. These chemostats are composed of four main parts: a syringe unit, a glass bottle (i.e. the chemostat), a medium bottle and a waste bottle, all connected by tubing. A peristaltic pump and a low overpressure in the system allow the flow of medium from the reservoir to the chemostat bottle and of unused nutrients, waste products and organisms to the waste bottle. Chemostats are put on stirring plates to create a homogenosus environment within.Becks, L., Hilker, F.M., Malchow, H., Jürgens, K., Arndt, H., 2005. Experimental demonstration of chaos in a microbial food web. Nature 435, 1226–1229. https://doi.org/10.1038/nature03627Boer, V.M., Crutchfield, C.A., Bradley, P.H., Botstein, D., Rabinowitz, J.D., 2010. Growth-limiting Intracellular Metabolites in Yeast Growing under Diverse Nutrient Limitations 21, 198–211. https://doi.org/10.1091/mbc.E09Frickel, J., Sieber, M., Becks, L., 2016. Eco-evolutionary dynamics in a coevolving host – virus system. Ecol. Lett. 19, 450–459. https://doi.org/10.1111/ele.12580Smith, H., Waltman, P., 1995.The Theory of the Chemostat: Dynamics of Microbial Competition (Cambridge Studies in Mathematical Biology). Cambridge: Cambridge University Press. doi:10.1017/CBO9780511530043Weitz, J.S. 2015. Quantitative Viral Ecology. Dynamics of viruses and their microbial host. Princeton University Press. ISBN: 9780691161549 .justify:after { content: ""; display:inline-block; width: 100%; }

Proper citation: Ana Del Arco, Noemi Woltermann, Lutz Becks 2020. Building up chemostats for experimental eco-evolutionary studies. protocols.io https://dx.doi.org/10.17504/protocols.io.tkxekxn Copy   

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Authors: B.R. Cunningham, J.R. Brum, S.M. Schwenck, M.B. Sullivan, S.G. John
Group: VERVE Net, Sullivan Lab
Summary: For use in Wet-Mount Method for Enumeration of Aquatic Viruses

Proper citation: B.R. Cunningham, J.R. Brum, S.M. Schwenck, M.B. Sullivan, S.G. John 2016. Iron Chloride Solution. protocols.io https://dx.doi.org/10.17504/protocols.io.c8wzxd Copy   

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Authors: Mathias Middelboe, Amy M. Chan, and Sif K. Bertelsen
Group: VERVE Net, Suttle Laboratory of Marine Molecular Microbiology and Virology
Summary: The life cycle of phages can be characterized by one-step growth experiments, which are designed in a way that allows only a single infection cycle to take place (i.e., no re-infections occurring by phages produced during the experiment).

Proper citation: Mathias Middelboe, Amy M. Chan, and Sif K. Bertelsen 2016. One-step growth experiments (bacteriophages). protocols.io https://dx.doi.org/10.17504/protocols.io.dpw5pd Copy   

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Authors: Simon Roux
Group: VERVE Net, Hurwitz Lab
Summary: METAVIR is a web server designed to annotate viral metagenomic sequences (raw reads or assembled contigs). A set of published viromes, identified as "public projects", is already available, and your own data sets can be processed in a private environment.

Proper citation: Simon Roux 2016. MetaVir- Analysis of viromes. protocols.io https://dx.doi.org/10.17504/protocols.io.d5i84d Copy   

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Authors: Matthew Sullivan
Group: VERVE Net, Sullivan Lab
Summary: This mixture is used for the DNA Extraction Protocol.

Proper citation: Matthew Sullivan 2016. CsCl purified phage lysate, Proteinase K, SDS. protocols.io https://dx.doi.org/10.17504/protocols.io.c68zhv Copy   

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Authors: Corina P. D. Brussaard, Jérôme P. Payet, Christian Winter, and Markus G. Weinbauer
Group: VERVE Net
Summary: This is a protocol from: Brussaard, C. P. D., J. P. Payet, C. Winter, and M. G. Weinbauer. 2010. Chapter 11: Quantification of aquatic viruses by flow cytometry. Manual of Aquatic Viral Ecology. Waco, TX:American Society of Limnology and Oceanography. doi:10.4319/mave.2010.978-0-9845591-0-7Please see the published manuscript for additional information.

Proper citation: Corina P. D. Brussaard, Jérôme P. Payet, Christian Winter, and Markus G. Weinbauer 2016. Quantification of aquatic viruses by flow cytometry. protocols.io https://dx.doi.org/10.17504/protocols.io.dpj5km Copy   

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Authors: David Dunigan and Irina Agarkova
Group: VERVE Net

Proper citation: David Dunigan and Irina Agarkova 2016. Transformation of Bacterial Cultures Using The Calcium Chloride Procedure. protocols.io https://dx.doi.org/10.17504/protocols.io.erbbd2n Copy   

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Authors: HANNIGAN GD, GRICE EA, ET AL.
Group: VERVE Net, Club Grice
Summary: This protocol provides a method for identifying bacteriophage/virus taxonomy without a virome dataset using UniProt reference database. Based on the methods found in the following publication:Hannigan, Geoffrey D., et al. "The Human Skin Double-Stranded DNA Virome: Topographical and Temporal Diversity, Genetic Enrichment, and Dynamic Associations with the Host Microbiome." mBio 6.5 (2015): e01578-15.

Proper citation: HANNIGAN GD, GRICE EA, ET AL. 2016. Script P4: Assigning Viral Taxonomy. protocols.io https://dx.doi.org/10.17504/protocols.io.efqbbmw Copy   

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Authors: Matthew Sullivan
Group: VERVE Net, Sullivan Lab

Proper citation: Matthew Sullivan 2016. DNA/RNA Extractions from Sterivex Filters. protocols.io https://dx.doi.org/10.17504/protocols.io.c4bysm Copy   

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Authors: HANNIGAN GD, GRICE EA, ET AL.
Group: VERVE Net, Club Grice
Summary: This protocol outlines the analysis used to plot KEGG functional annotation.

Proper citation: HANNIGAN GD, GRICE EA, ET AL. 2016. Script R12: Functional Analysis. protocols.io https://dx.doi.org/10.17504/protocols.io.ejibcke Copy   

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