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CyanoGate: A modular cloning suite for engineering cyanobacteria based on the plant MoClo syntax

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    Rights statement: Copyright © 2019 American Society of Plant Biologists. All rights reserved. Copyright 2019 by the American Society of Plant Biologists.

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Original languageEnglish
Pages (from-to)39-55
Number of pages17
JournalPlant physiology
Volume180
Issue number1
Early online date28 Feb 2019
DOIs
Publication statusE-pub ahead of print - 28 Feb 2019

Abstract

Recent advances in synthetic biology research have been underpinned by an exponential increase in available genomic information and a proliferation of advanced DNA assembly tools. The adoption of plasmid vector assembly standards and parts libraries has greatly enhanced the reproducibility of research and exchange of parts between different labs and biological systems. However, a standardised Modular Cloning (MoClo) system is not yet available for cyanobacteria, which lag behind other prokaryotes in synthetic biology despite their huge potential in biotechnological applications. By building on the assembly library and syntax of the Plant Golden Gate MoClo kit, we have developed a versatile system called CyanoGate that unites cyanobacteria with plant and algal systems. Here we have generated a suite of parts and acceptor vectors for making i) marked/unmarked knock-outs or integrations using an integrative acceptor vector, and ii) transient multigene expression and repression systems using known and novel replicative vectors. We have tested and compared the CyanoGate system in the established model cyanobacterium Synechocystis sp. PCC 6803 and the more recently described fast-growing strain Synechococcus elongatus UTEX 2973. We observed that fast-growth phenotype in UTEX 2973 is only evident under specific growth conditions, but that UTEX 2973 can accumulate high levels of proteins with strong native or synthetic promoters. The system is publicly available and can be readily expanded to accommodate other standardised MoClo parts to accelerate the development of reliable synthetic biology tools for the cyanobacterial community.

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