ArticleBMC biotechnology2018
Agrobacterium-mediated and electroporation-mediated transformation of Chlamydomonas reinhardtii: a comparative study.
Article in BMC biotechnology, 2018. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 13 papers.
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Who cites it
13 citing papers in PubMed, 42 citations in OpenAlex.
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- Cell-Penetrating Peptide-Based Triple Nanocomplex Enables Efficient Nuclear Gene Delivery in Chlamydomonas reinhardtii.Biotechnology and bioengineering · 2025Article
- Genome editing in the green alga Chlamydomonas: past, present practice and future prospects.The Plant journal : for cell and molecular biology · 2025Review
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- Current Status and Perspective on the Use of Viral-Based Vectors in Eukaryotic Microalgae.Marine drugs · 2022Review
- Efficient3 Biotech · 2021Article
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- Genome Editing by CRISPR-Cas: A Game Change in the Genetic Manipulation ofLife (Basel, Switzerland) · 2020Review
- Bioengineering of Microalgae: Recent Advances, Perspectives, and Regulatory Challenges for Industrial Application.Frontiers in bioengineering and biotechnology · 2020Review
- When Unity Is Strength: The Strategies Used byCells · 2019Review
- Simplified Transformation ofGenes · 2019Article
- Evaluation of strategies for improving the transgene expression in an oleaginous microalga Scenedesmus acutus.BMC biotechnology · 2019Article
- Biomass from microalgae: the potential of domestication towards sustainable biofactories.Microbial cell factories · 2018Review
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Authors and funding
7 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundChlamydomonas reinhardtii is an unicellular green alga used for functional genomics studies and heterologous protein expression. A major hindrance in these studies is the low level and instability of expression of nuclear transgenes, due to their rearrangement and/or silencing over time.
resultsWe constructed dedicated vectors for Agrobacterium-mediated transformation carrying, within the T-DNA borders, the Paromomycin (Paro) selectable marker and an expression cassette containing the Luciferase (Luc) reporter gene. These vectors and newly developed co-cultivation methods were used to compare the efficiency, stability and insertion sites of Agrobacterium- versus electroporation-mediated transformation. The influence of different transformation methods, of the cell wall, of the virulence of different Agrobacterium strains, and of transgene orientation with respect to T-DNA borders were assessed. False positive transformants were more frequent in Agrobacterium-mediated transformation compared to electroporation, compensating for the slightly lower proportion of silenced transformants observed in Agrobacterium-mediated transformation than in electroporation. The proportion of silenced transformants remained stable after 20 cycles of subculture in selective medium. Next generation sequencing confirmed the nuclear insertion points, which occurred in exons or untraslated regions (UTRs) for 10 out of 10 Agrobacterium-mediated and 9 out of 13 of electroporation-mediated insertions. Electroporation also resulted in higher numbers of insertions at multiple loci.
conclusionsDue to its labor-intensive nature, Agrobacterium transformation of Chlamydomonas does not present significant advantages over electroporation, with the possible exception of its use in insertional mutagenesis, due to the higher proportion of within-gene, single-locus insertions. Our data indirectly support the hypothesis that rearrangement of transforming DNA occurs in the Chlamydomonas cell, rather than in the extracellular space as previously proposed.
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