Cloning and expression of Burkholderia polyyne biosynthetic gene clusters in 2 Paraburkholderia hosts provides a strategy for biopesticide development

Petrova, Yoana D., Zhao, Jinlian, Webster, Gordon ORCID: https://orcid.org/0000-0002-9530-7835, Mullins, Alex J. ORCID: https://orcid.org/0000-0001-5804-9008, Williams, Katherine, Alswat, Amal S, Challis, Gregory L., Bailey, Andy and Mahenthiralingam, Eshwar ORCID: https://orcid.org/0000-0001-9014-3790 2022. Cloning and expression of Burkholderia polyyne biosynthetic gene clusters in 2 Paraburkholderia hosts provides a strategy for biopesticide development. Microbial Biotechnology 15 (10) , pp. 2547-2561. 10.1111/1751-7915.14106

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Abstract

Burkholderia have potential as biocontrol agents because they encode diverse biosynthetic gene clusters (BGCs) for a range of antimicrobial metabolites. Given the opportunistic pathogenicity associated with Burkholderia species, heterologous BGC expression within non-pathogenic hosts is a strategy to construct safe biocontrol strains. We constructed a yeast-adapted Burkholderia-Escherichia shuttle vector (pMLBAD_yeast) with a yeast replication origin 2 μ and URA3 selection marker and optimised it for cloning BGCs using the in vivo recombination ability of Saccharomyces cerevisiae. Two Burkholderia polyyne BGCs, cepacin (13 kb) and caryoynencin (11 kb), were PCR-amplified as three overlapping fragments, cloned downstream of the pBAD arabinose promoter in pMLBAD_yeast and mobilised into Burkholderia and Paraburkholderia heterologous hosts. Paraburkholderia phytofirmans carrying the heterologous polyyne constructs displayed in vitro bioactivity against a variety of fungal and bacterial plant pathogens similar to the native polyyne producers. Thirteen Paraburkholderia strains with preferential growth at 30°C compared with 37°C were also identified, and four of these were amenable to genetic manipulation and heterologous expression of the caryoynencin construct. The cloning and successful heterologous expression of Burkholderia biosynthetic gene clusters within Paraburkholderia with restricted growth at 37°C opens avenues for engineering non-pathogenic biocontrol strains.

Item Type:	Article
Date Type:	Publication
Status:	Published
Schools:	Biosciences
Additional Information:	This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
Publisher:	Wiley
ISSN:	1751-7915
Funders:	Biotechnology and Biological Sciences Research Council
Date of First Compliant Deposit:	17 June 2022
Date of Acceptance:	9 June 2022
Last Modified:	29 Mar 2024 05:31
URI:	https://orca.cardiff.ac.uk/id/eprint/150580

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