Journal
ALGAL RESEARCH-BIOMASS BIOFUELS AND BIOPRODUCTS
Volume 33, Issue -, Pages 448-455Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.algal.2018.06.021
Keywords
Electroporation; Green microalgae; Labelled DNA; Labelled protein; Sytox Green; Propidium iodide
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Funding
- Food and Nutrition Delta program of Agentschap NL [FND10007]
- Unilever
- National Commission of Scientific and Technologic Research of Chile (CONICYT)
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Genetic transformation of microalgae remains a challenge due to poor intracellular delivery of exogenous molecules. This limitation is caused by the structure and composition of the cell wall and cell membrane of each species. Moreover, successful delivery of proteins or nucleic acids cannot be assessed by determining trans-formability since their functionality is not always known in the studied microorganism. We propose a quick and effective screening tool for the prediction and optimization of electroporation settings by monitoring cell permeability and viability using Sytox Green and propidium iodide respectively. We determined voltage settings for the microalgae Chlamydomonas reinhardtii, Chlorella vulgaris, Neochloris oleoabundans and Acutodesmus obliquus. To evaluate the predicted settings, we delivered labelled DNA and proteins into the cells. We demonstrated that high transformation efficiencies can be accomplished when predicted values were applied with functional plasmids. Additionally, we increased transformation efficiencies by testing cell concentrations, light intensities and fragment sizes. This method can be used to determine suitable transformation conditions for non-transformed microalgae species and to increase the insight on established transformation protocols.
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