Stable transformation of the green algae Acutodesmus obliquus and Neochloris oleoabundans based on E. coli conjugation

Muñoz, Camilo F.; Sturme, Mark H. J.; D’Adamo, Sarah; Weusthuis, Ruud A.; Wijffels, René H.

doi:10.1016/j.algal.2019.101453

Cited by 27 publications

(18 citation statements)

References 34 publications

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“…Moreover, the elimination of episomal CRISPR/Cas9 vector from the host upon removal of selection pressure makes the mutant lines be considered as non-transgenic. In contrast to diatom, using similar vector system, recently Muñoz et al ( 2019 ) were not able to rescue the episomal plasmids from positive transformants of green oleaginous microalgae Acutodesmus obliquus and Neochloris oleoabundans . Moreover, the continuous subculturing in the selection-free medium was not sufficient to remove the episomal vector.…”

Section: Advancement In the Resources For Microalgal Researchmentioning

confidence: 97%

“…The plausible explanation for this delayed mutant appearance was attributed to the lower Cas9 expression due to higher rate of cell division in conjugatively transformed cells. In addition, the episomal vector system adapted for diatom was able to transform the green oleaginous microalgae Acutodesmus obliquus and Neochloris oleoabundans through bacterial conjugation (Muñoz et al, 2019 ). Although the transformation efficiency was sufficiently higher as compared to the biolistic-based vector delivery system, this application of diatom adapted episomal vector system in other microalgae has some limitations that are discussed in the following section.…”

Section: Advancement In the Resources For Microalgal Researchmentioning

confidence: 99%

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Bioengineering of Microalgae: Recent Advances, Perspectives, and Regulatory Challenges for Industrial Application

Kumar

Shekh

Jakhu

et al. 2020

Front. Bioeng. Biotechnol.

194

123

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Microalgae, due to their complex metabolic capacity, are being continuously explored for nutraceuticals, pharmaceuticals, and other industrially important bioactives. However, suboptimal yield and productivity of the bioactive of interest in local and robust wild-type strains are of perennial concerns for their industrial applications. To overcome such limitations, strain improvement through genetic engineering could play a decisive role. Though the advanced tools for genetic engineering have emerged at a greater pace, they still remain underused for microalgae as compared to other microorganisms. Pertaining to this, we reviewed the progress made so far in the development of molecular tools and techniques, and their deployment for microalgae strain improvement through genetic engineering. The recent availability of genome sequences and other omics datasets form diverse microalgae species have remarkable potential to guide strategic momentum in microalgae strain improvement program. This review focuses on the recent and significant improvements in the omics resources, mutant libraries, and high throughput screening methodologies helpful to augment research in the model and non-model microalgae. Authors have also summarized the case studies on genetically engineered microalgae and highlight the opportunities and challenges that are emerging from the current progress in the application of genome-editing to facilitate microalgal strain improvement. Toward the end, the regulatory and biosafety issues in the use of genetically engineered microalgae in commercial applications are described.

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Section: Advancement In the Resources For Microalgal Researchmentioning

confidence: 97%

Section: Advancement In the Resources For Microalgal Researchmentioning

confidence: 99%

Bioengineering of Microalgae: Recent Advances, Perspectives, and Regulatory Challenges for Industrial Application

Kumar

Shekh

Jakhu

et al. 2020

Front. Bioeng. Biotechnol.

194

123

View full text Add to dashboard Cite

show abstract

“…Bacterial conjugation has been used commonly in cyanobacterial species such as Synechococcus, Prochlorococcus, N. punctiforme, Anabaena , and Synechocystis sp. and recently in eukaryotic algae such as P. tricornutum, Thalassiosira pseudonana, Acutodesmus obliquus, and Neochloris oleoabundans [ 79 , 81 , 82 , 83 , 84 , 85 , 86 , 87 , 88 ]. The effectiveness of transformation by conjugation is based on the capability of the algal recipient strain to integrate and maintain the vector, either in the chromosome, in endogenous plasmids, or as an episomal plasmid [ 78 , 79 , 87 , 88 ].…”

Section: Natural Transformation Bacterial Conjugation and Agrobacterium -Mediated Transformationmentioning

confidence: 99%

“…Conjugation has the advantage over electroporation, microparticle bombardment, and glass-beads agitation to deliver larger DNA fragments [ 45 ]. The use of the conjugation-based method in P. tricornutum, N. oceanica, and Neochloris oleoabundans has been demonstrated to generate mutants with higher transformation efficiency compared to microparticle bombardment transformation: 500–1000 transformants/10 8 cells/200 ng DNA (conjugation) compared to 5–25 transformants/10 8 cells/2.5 µg DNA (bombardment) in P. tricornutum [ 87 , 94 , 95 , 96 ].…”

Section: Natural Transformation Bacterial Conjugation and Agrobacterium -Mediated Transformationmentioning

confidence: 99%

Gene Delivery Technologies with Applications in Microalgal Genetic Engineering

Gutiérrez

Lauersen

2021

Biology

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Microalgae and cyanobacteria are photosynthetic microbes that can be grown with the simple inputs of water, carbon dioxide, (sun)light, and trace elements. Their engineering holds the promise of tailored bio-molecule production using sustainable, environmentally friendly waste carbon inputs. Although algal engineering examples are beginning to show maturity, severe limitations remain in the transformation of multigene expression cassettes into model species and DNA delivery into non-model hosts. This review highlights common and emerging DNA delivery methods used for other organisms that may find future applications in algal engineering.

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“…Truncated sgRNAs with 17 to 19-nt spacer are more sensitive to mismatches than others with 20-nt length, which can effectively reduce off-target mutations. Besides, the recent study [ 60 ] has shown that shorter spacers such as 17 to 18 nt have higher specificity but perform less on-target effective than 19 or 20-nt spacers. Thus, the choice here is tough to design the best sgRNA guide sequence without creating mismatches.…”

Section: Crispr-cas System In C Reinhardtiimentioning

confidence: 99%

Genome Editing by CRISPR-Cas: A Game Change in the Genetic Manipulation of Chlamydomonas

Ghribi

Nouemssi

Meddeb-Mouelhi

et al. 2020

Life

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Microalgae are promising photosynthetic unicellular eukaryotes among the most abundant on the planet and are considered as alternative sustainable resources for various industrial applications. Chlamydomonas is an emerging model for microalgae to be manipulated by multiple biotechnological tools in order to produce high-value bioproducts such as biofuels, bioactive peptides, pigments, nutraceuticals, and medicines. Specifically, Chlamydomonas reinhardtii has become a subject of different genetic-editing techniques adapted to modulate the production of microalgal metabolites. The main nuclear genome-editing tools available today include zinc finger nucleases (ZFNs), transcriptional activator-like effector nucleases (TALENs), and more recently discovered the clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR associated protein (Cas) nuclease system. The latter, shown to have an interesting editing capacity, has become an essential tool for genome editing. In this review, we highlight the available literature on the methods and the applications of CRISPR-Cas for C. reinhardtii genetic engineering, including recent transformation methods, most used bioinformatic tools, best strategies for the expression of Cas protein and sgRNA, the CRISPR-Cas mediated gene knock-in/knock-out strategies, and finally the literature related to CRISPR expression and modification approaches.

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Stable transformation of the green algae Acutodesmus obliquus and Neochloris oleoabundans based on E. coli conjugation

Cited by 27 publications

References 34 publications

Bioengineering of Microalgae: Recent Advances, Perspectives, and Regulatory Challenges for Industrial Application

Bioengineering of Microalgae: Recent Advances, Perspectives, and Regulatory Challenges for Industrial Application

Gene Delivery Technologies with Applications in Microalgal Genetic Engineering

Genome Editing by CRISPR-Cas: A Game Change in the Genetic Manipulation of Chlamydomonas

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