CRISPR-Cas9 Shaped Viral Metagenomes Associated with Bacillus subtilis

Katharina Kohm; Veronika Theresa Lutz; Ines Friedrich; Robert Hertel

doi:10.1007/978-1-0716-2795-2_15

CRISPR-Cas9 Shaped Viral Metagenomes Associated with Bacillus subtilis

Methods Mol Biol. 2023:2555:205-212. doi: 10.1007/978-1-0716-2795-2_15.

Authors

Katharina Kohm¹, Veronika Theresa Lutz², Ines Friedrich³, Robert Hertel^{4

5}

Affiliations

¹ Institute of Biotechnology, BTU Cottbus-Senftenberg, Senftenberg, Germany.
² Department of Veterinary and Animal Sciences, University of Copenhagen, Frederiksberg, Denmark.
³ Institute of Microbiology and Genetics, Georg August University Göttingen, Göttingen, Germany. ines.friedrich@uni-goettingen.de.
⁴ Institute of Biotechnology, BTU Cottbus-Senftenberg, Senftenberg, Germany. rhertel@gwdg.de.
⁵ Institute of Microbiology and Genetics, Georg August University Göttingen, Göttingen, Germany. rhertel@gwdg.de.

PMID: 36306089
DOI: 10.1007/978-1-0716-2795-2_15

Abstract

Phages are viruses of bacteria and have been known for over a century. They do not have a metabolism or protein synthesis machinery and rely on host cells for replication. The model organism Bacillus subtilis has served as a host strain for decades and enabled the isolation of many unique viral strains. However, many viral species representatives remained orphans as no, or only a few, related phages were ever re-isolated.The presented protocol describes how a CRISPR-Cas9 system with an artificial CRISPR-array can be set up and used to discriminate abundant and well-known B. subtilis phage from a host-based metagenome enrichment. The obtained viral suspension can be used for metagenome sequencing and isolating new viral strains.

Keywords: Bacillus subtilis; Bacteriophage; CRISPR-Cas9; CutSPR; Metagenome; Phage; gRNA.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Bacillus subtilis* / genetics
Bacteriophages*
CRISPR-Cas Systems / genetics
Metagenome