Development of CRISPR/Cas9 system in Chlorella vulgaris FSP-E to enhance lipid accumulation

[Display omitted] •Using LB and RB homologous flanks led to successful Chlorella transformation.•GFP expression in transformants was observed by spectrophotometer and microscopy.•The CRISPR/Cas9 system was the first time to apply in Chlorella vulgaris FSP-E.•Targeting fad3 gene by CRISPR technology...

Full description

Saved in:
Bibliographic Details
Published in:Enzyme and microbial technology 2020-02, Vol.133, p.109458-109458, Article 109458
Main Authors: Lin, Way-Rong, Ng, I-Son
Format: Article
Language:English
Subjects:
Agrobacterium tumefaciens - genetics
Biofuels
Biomass
Biorefinery
Chlorella vulgaris
Chlorella vulgaris - genetics
Chlorella vulgaris - metabolism
CRISPR-Cas Systems
CRISPR/Cas9
Fatty acid desaturase
Fatty Acid Desaturases - genetics
Green Fluorescent Proteins - genetics
Lipid
Lipid Metabolism
Lipids - analysis
Microalgae - genetics
Microalgae - metabolism
Plasmids - genetics
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:[Display omitted] •Using LB and RB homologous flanks led to successful Chlorella transformation.•GFP expression in transformants was observed by spectrophotometer and microscopy.•The CRISPR/Cas9 system was the first time to apply in Chlorella vulgaris FSP-E.•Targeting fad3 gene by CRISPR technology improved lipid content of Chlorella. Microalgae biorefinery is an alternative, sustainable and promising trend to solve the problem of fossil oil depletion and carbon dioxide emission. However, considering the innate limitation of cell growth and oil content in microalgae, to accelerate metabolic balance by CRISPR/Cas9 system is attractive. At first, plasmid based from Agrobacterium tumefaciens and a fragment of mGFP was transformed into Chlorella sorokiniana and Chlorella vulgaris FSP-E by electroporation, respectively. Selected colonies were tested by spectrophotometer and inverted fluorescence microscopy (IFM), and an increase of fluorescent was observed by 67% compared with that in wild type, which proved the Agrobacterium-mediated plasmid is suitable for gene insertion in Chlorella species. Consequently, plasmid with similar structure as mentioned previously containing fragment of Cas9 with sgRNA designed on omega-3 fatty acid desaturase (fad3) gene was constructed and showed a higher accumulation of lipid content by 46% (w/w) in C. vulgaris FSP-E. This is first-time to use CRISPR/Cas9 based technology for gene manipulation in Chlorella.
ISSN:0141-0229
1879-0909
DOI:10.1016/j.enzmictec.2019.109458