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Development of a forward genetic screen to isolate oil mutants in the green microalga Chlamydomonas reinhardtii
- Source :
- Biotechnology for Biofuels, Biotechnology for Biofuels, BioMed Central, 2013, 6 (1), ⟨10.1186/1754-6834-6-178⟩, Biotechnology for Biofuels, 2013, 6 (1), ⟨10.1186/1754-6834-6-178⟩
- Publication Year :
- 2013
- Publisher :
- HAL CCSD, 2013.
-
Abstract
- International audience; Oils produced by microalgae are precursors to biodiesel. To achieve a profitable production of biodiesel from microalgae, identification of factors governing oil synthesis and turnover is desirable. The green microalga Chlamydomonas reinhardtii is amenable to genetic analyses and has recently emerged as a model to study oil metabolism. However, a detailed method to isolate various types of oil mutants that is adapted to Chlamydomonas has not been reported.ResultsWe describe here a forward genetic approach to isolate mutants altered in oil synthesis and turnover from C. reinhardtii. It consists of a three-step screening procedure: a primary screen by flow cytometry of Nile red stained transformants grown in 96-deep-well plates under three sequential conditions (presence of nitrogen, then absence of nitrogen, followed by oil remobilization); a confirmation step using Nile red stained biological triplicates; and a validation step consisting of the quantification by thin layer chromatography of oil content of selected strains. Thirty-one mutants were isolated by screening 1,800 transformants generated by random insertional mutagenesis (1.7%). Five showed increased oil accumulation under the nitrogen-replete condition and 13 had altered oil content under nitrogen-depletion. All mutants were affected in oil remobilization.ConclusionThis study demonstrates that various types of oil mutants can be isolated in Chlamydomonas based on the method set-up here, including mutants accumulating oil under optimal biomass growth. The strategy conceived and the protocol set-up should be applicable to other microalgal species such as Nannochloropsis and Chlorella, thus serving as a useful tool in Chlamydomonas oil research and algal biotechnology.
- Subjects :
- 0106 biological sciences
[SDV]Life Sciences [q-bio]
Mutant
Mutagenesis (molecular biology technique)
Chlamydomonas reinhardtii
Management, Monitoring, Policy and Law
Biology
Genetic screen
01 natural sciences
Applied Microbiology and Biotechnology
03 medical and health sciences
Bioenergy
Botany
Chlamydomonas mutants
Food science
Flow cytometry
030304 developmental biology
2. Zero hunger
0303 health sciences
Biodiesel
Microalgal oil
Renewable Energy, Sustainability and the Environment
Lipid remobilization
Chlamydomonas
Methodology
Nile red
biology.organism_classification
General Energy
Biofuel
010606 plant biology & botany
Biotechnology
Subjects
Details
- Language :
- English
- ISSN :
- 17546834
- Database :
- OpenAIRE
- Journal :
- Biotechnology for Biofuels, Biotechnology for Biofuels, BioMed Central, 2013, 6 (1), ⟨10.1186/1754-6834-6-178⟩, Biotechnology for Biofuels, 2013, 6 (1), ⟨10.1186/1754-6834-6-178⟩
- Accession number :
- edsair.doi.dedup.....300c1a0a23ff79f9ba37e3e9d4e177e0