A toolbox to engineer the highly productive cyanobacterium Synechococcus sp. PCC 11901.

Autor:	Victoria AJ; Institute of Molecular Plant Sciences, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK.; Centre for Engineering Biology, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK., Selão TT; Department of Chemical and Environmental Engineering, University of Nottingham, Nottingham NG7 2RD, UK., Moreno-Cabezuelo JÁ; School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, UK., Mills LA; School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, UK., Gale GAR; Institute of Molecular Plant Sciences, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK.; Centre for Engineering Biology, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK., Lea-Smith DJ; School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, UK., McCormick AJ; Institute of Molecular Plant Sciences, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK.; Centre for Engineering Biology, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3BF, UK.
Jazyk:	angličtina
Zdroj:	Plant physiology [Plant Physiol] 2024 Oct 01; Vol. 196 (2), pp. 1674-1690.
DOI:	10.1093/plphys/kiae261
Abstrakt:	Synechococcus sp. PCC 11901 (PCC 11901) is a fast-growing marine cyanobacterial strain that has a capacity for sustained biomass accumulation to very high cell densities, comparable to that achieved by commercially relevant heterotrophic organisms. However, genetic tools to engineer PCC 11901 for biotechnology applications are limited. Here we describe a suite of tools based on the CyanoGate MoClo system to unlock the engineering potential of PCC 11901. First, we characterized neutral sites suitable for stable genomic integration that do not affect growth even at high cell densities. Second, we tested a suite of constitutive promoters, terminators, and inducible promoters including a 2,4-diacetylphloroglucinol (DAPG)-inducible PhlF repressor system, which has not previously been demonstrated in cyanobacteria and showed tight regulation and a 228-fold dynamic range of induction. Lastly, we developed a DAPG-inducible dCas9-based CRISPR interference (CRISPRi) system and a modular method to generate markerless mutants using CRISPR-Cas12a. Based on our findings, PCC 11901 is highly responsive to CRISPRi-based repression and showed high efficiencies for single insertion (31% to 81%) and multiplex double insertion (25%) genome editing with Cas12a. We envision that these tools will lay the foundations for the adoption of PCC 11901 as a robust model strain for engineering biology and green biotechnology. Competing Interests: Conflict of interest statement. None declared. (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society of Plant Biologists.)
Databáze:	MEDLINE
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