|Title||Improved DNA/protein delivery in microalgae – A simple and reliable method for the prediction of optimal electroporation settings|
|Author(s)||Muñoz, Camilo F.; Jaeger, Lenny de; Sturme, Mark H.J.; Lip, Ka Y.F.; Olijslager, Justus W.J.; Springer, Jan; Wolbert, Emil J.H.; Martens, Dirk E.; Eggink, Gerrit; Weusthuis, Ruud A.; Wijffels, René H.|
|Source||Algal Research 33 (2018). - ISSN 2211-9264 - p. 448 - 455.|
|Publication type||Refereed Article in a scientific journal|
|Keyword(s)||Electroporation - Green microalgae - Labelled DNA - Labelled protein - Propidium iodide - Sytox Green|
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 transformability 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.