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Methods Mol Biol. 2015;1311:223-32. doi: 10.1007/978-1-4939-2687-9_14.

Archaeal Viruses of the Sulfolobales: Isolation, Infection, and CRISPR Spacer Acquisition.

Author information

1
Archaea Centre, Department of Biology, University of Copenhagen, Ole Maal√łes Vej 5, 2200, Copenhagen N, Denmark, s.erdmann@unsw.edu.au.

Abstract

Infection of archaea with phylogenetically diverse single viruses, performed in different laboratories, has failed to activate spacer acquisition into host CRISPR loci. The first successful uptake of archaeal de novo spacers was observed on infection of Sulfolobus solfataricus P2 with an environmental virus mixture isolated from Yellowstone National Park (Erdmann and Garrett, Mol Microbiol 85:1044-1056, 2012). Experimental studies of isolated genetic elements from this mixture revealed that SMV1 (S ulfolobus Monocauda Virus 1), a tailed spindle-shaped virus, can induce spacer acquisition in CRISPR loci of Sulfolobus species from a second coinfecting conjugative plasmid or virus (Erdmann and Garrett, Mol Microbiol 85:1044-1056, 2012; Erdmann et al. Mol Microbiol 91:900-917, 2014). Here we describe, firstly, the isolation of archaeal virus mixtures from terrestrial hot springs and the techniques used both to infect laboratory strains with these virus mixtures and to obtain purified virus particles. Secondly, we present the experimental conditions required for activating SMV1-induced spacer acquisition in two different Sulfolobus species.

PMID:
25981476
DOI:
10.1007/978-1-4939-2687-9_14
[Indexed for MEDLINE]

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