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Lea-Smith, David J. 
ORCID: https://orcid.org/0000-0003-2463-406X, Vasudevan, Ravendran and Howe, Christopher J.
(2016)
Generation of marked and markerless mutants in model cyanobacterial species.
Journal of Visualized Experiments, 2016 (111).
ISSN 1940-087X
Abstract
Cyanobacteria are ecologically important organisms and potential platforms for production of biofuels and useful industrial products. Genetic manipulation of cyanobacteria, especially model organisms such as Synechocystis sp. PCC6803 and Synechococcus sp. PCC7002, is a key tool for both basic and applied research. Generation of unmarked mutants, whereby chromosomal alterations are introduced into a strain via insertion of an antibiotic resistance cassette (a manipulatable fragment of DNA containing one or more genes), followed by subsequent removal of this cassette using a negative selectable marker, is a particularly powerful technique. Unmarked mutants can be repeatedly genetically manipulated, allowing as many alterations to be introduced into a strain as desired. In addition, the absence of genes encoding antibiotic resistance proteins in the mutated strain is desirable, as it avoids the possibility of 'escape' of antibiotic resistant organisms into the environment. However, detailed methods for repeated rounds of genetic manipulation of cyanobacteria are not well described in the scientific literature. Here we provide a comprehensive description of this technique, which we have successfully used to generate mutants with multiple deletions, single point mutations within a gene of interest and insertion of novel gene cassettes.
| Item Type: | Article |
|---|---|
| Additional Information: | Publisher Copyright: © 2016 Journal of Visualized Experiments. |
| Uncontrolled Keywords: | biofuels,cyanobacteria,issue 111,microbiology,natural products,photosynthesis,sacb,synechococcus,synechocystis,unmarked mutants,general neuroscience,general chemical engineering,general biochemistry,genetics and molecular biology,general immunology and microbiology ,/dk/atira/pure/subjectarea/asjc/2800/2800 |
| Faculty \ School: | University of East Anglia Research Groups/Centres > Theme - ClimateUEA Faculty of Science > School of Biological Sciences |
| UEA Research Groups: | Faculty of Science > Research Groups > Molecular Microbiology |
| Related URLs: | |
| Depositing User: | LivePure Connector |
| Date Deposited: | 01 Jul 2026 11:56 |
| Last Modified: | 01 Jul 2026 13:05 |
| URI: | https://ueaeprints.uea.ac.uk/id/eprint/103583 |
| DOI: | 10.3791/54001 |
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