Journal article
Comprehensive Genome Engineering Toolbox for Microalgae Nannochloropsis oceanica Based on CRISPR-Cas Systems
ACS synthetic biology, Vol.10(12), pp.3369-3378
12/17/2021
DOI: 10.1021/acssynbio.1c00329
PMCID: PMC8689688
PMID: 34793143
Abstract
Microalgae can produce industrially relevant metabolites using atmospheric CO
and sunlight as carbon and energy sources, respectively. Developing molecular tools for high-throughput genome engineering could accelerate the generation of tailored strains with improved traits. To this end, we developed a genome editing strategy based on Cas12a ribonucleoproteins (RNPs) and homology-directed repair (HDR) to generate scarless and markerless mutants of the microalga
. We also developed an episomal plasmid-based Cas12a system for efficiently introducing indels at the target site. Additionally, we exploited the ability of Cas12a to process an associated CRISPR array to perform multiplexed genome engineering. We efficiently targeted three sites in the host genome in a single transformation, thereby making a major step toward high-throughput genome engineering in microalgae. Furthermore, a CRISPR interference (CRISPRi) tool based on Cas9 and Cas12a was developed for effective downregulation of target genes. We observed up to 85% reduction in the transcript levels upon performing CRISPRi with dCas9 in
. Overall, these developments substantially accelerate genome engineering efforts in
and potentially provide a general toolbox for improving other microalgal strains.
Details
- Title: Subtitle
- Comprehensive Genome Engineering Toolbox for Microalgae Nannochloropsis oceanica Based on CRISPR-Cas Systems
- Creators
- Mihris Ibnu Saleem Naduthodi - Wageningen University & ResearchChristian Südfeld - Wageningen University & ResearchEmmanouil Klimis Avitzigiannis - Wageningen University & ResearchNicola Trevisan - Wageningen University & ResearchEduard van Lith - Laboratory of Microbiology, Wageningen University, Wageningen 6708 WE, The NetherlandsJavier Alcaide Sancho - Laboratory of Microbiology, Wageningen University, Wageningen 6708 WE, The NetherlandsSarah D'Adamo - Wageningen University & ResearchMaria Barbosa - Wageningen University & ResearchJohn van der Oost - Wageningen University & Research
- Resource Type
- Journal article
- Publication Details
- ACS synthetic biology, Vol.10(12), pp.3369-3378
- DOI
- 10.1021/acssynbio.1c00329
- PMID
- 34793143
- PMCID
- PMC8689688
- NLM abbreviation
- ACS Synth Biol
- ISSN
- 2161-5063
- eISSN
- 2161-5063
- Grant note
- DOI: 10.13039/100010661, name: Horizon 2020 Framework Programme, award: 745754
- Language
- English
- Date published
- 12/17/2021
- Academic Unit
- International Programs; Spanish and Portuguese
- Record Identifier
- 9984398035702771
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