Genome integration and excision by a new Streptomyces Bacteriophage ϕJoe

Fogg, P. C.M., Haley, J. A., Stark, W. M. and Smith, M. C.M. (2017) Genome integration and excision by a new Streptomyces Bacteriophage ϕJoe. Applied and Environmental Microbiology, 83(5), e02767-16. (doi: 10.1128/AEM.02767-16) (PMID:28003200) (PMCID:PMC5311408)

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Abstract

Bacteriophages are the source of many valuable tools for molecular biology and genetic manipulation. In Streptomyces, most DNA cloning vectors are based on serine integrase site-specific DNA recombination systems derived from phage. Because of their efficiency and simplicity, serine integrases are also used for diverse synthetic biology applications. Here we present the genome of a new Streptomyces phage, ϕJoe, and investigate the conditions for integration and excision of the φJoe genome. ϕJoe belongs to the largest Streptomyces phage cluster (R4-like) and encodes a serine integrase. The attB site from S. venezuelae was used efficiently by an integrating plasmid, pCMF92, constructed using the ϕJoe int/attP locus. The attB site for ϕJoe integrase was occupied in several Streptomyces genomes, including S. coelicolor, by a mobile element that varies in gene content and size between host species. Serine integrases require a phage-encoded recombination directionality factor (RDF) to activate the excision reaction. The ϕJoe RDF was identified and its function was confirmed in vivo. Both the integrase and RDF were active in in vitro recombination assays. The ϕJoe site-specific recombination system is likely to be an important addition to the synthetic biology and genome engineering toolbox.

Item Type:Articles
Additional Information:This research was performed with funding from the Biotechnology and Biological Sciences Research Council (project grant BB/K003356/1) and the Microbiology Society (formerly Society for General Microbiology) with a Harry Smith Vacation Scholarship for JH.
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Stark, Professor Marshall
Authors: Fogg, P. C.M., Haley, J. A., Stark, W. M., and Smith, M. C.M.
College/School:College of Medical Veterinary and Life Sciences > Institute of Molecular Cell and Systems Biology
Journal Name:Applied and Environmental Microbiology
Publisher:American Society for Microbiology.
ISSN:0099-2240
ISSN (Online):1098-5336
Published Online:21 December 2016
Copyright Holders:Copyright © 2016 Fogg et al.
First Published:First published in Applied and Environmental Microbiology 83(5): e02767-16
Publisher Policy:Reproduced under a creative commons license

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
589141A platform for rapid and precise DNA module rearrangements in Synthetic BiologyWilliam StarkBiotechnology and Biological Sciences Research Council (BBSRC)BB/K003356/1RI MOLECULAR CELL & SYSTEMS BIOLOGY