Expanding the molecular toolbox for Zygnematophyceae - transient genetic transformation of the desmidMicrasterias radiansvar.evoluta

In the past, various species of the unicellular algal genusMicrasterias(Zygnematophyceae) have been used for research in the fields of cell biology and physiology. Relatively large cell size, highly differentiated cell shape and a remarkable evolutionary position makeMicrasteriasinteresting, especially for cytomorphogenetic studies. However, within this genus a model organism enabling molecular research has not yet been established.Micrasterias radiansvar.evoluta(W.B.Turner) Krieger allows easy culturing under axenic conditions and performs its whole life cyclein vitrothus fulfilling two important prerequisites for a model organism. In this work resources allowing transient expression of transgenes were developed. First, axenic lines were obtained by the treatment of zygospores with a cocktail of antibiotics followed by germination and regeneration. In order to allow transgene expression we isolated the 5 ' -flanking region of theM. radiansvar.evoluta Actin1gene (MrACT1) and fused it to the green fluorescent protein (GFP). A higher promoter activity compared with various heterologous promoters regarding GFP expression was observed. Transient transgene expression was achieved by polyethylene glycol (PEG)-mediated protoplast transformation, yielding a rate of 3.9% transformed cells per surviving protoplasts. Transgene expression was also achieved by particle bombardment of vegetative cells. Employing the established protocol, a Lifeact-GFP fusion protein for labelling F-actin was expressed, allowing visualization of the actin cytoskeleton inM. radiansvar.evoluta.

Automated summary

In this study, resources allowing transient expression of transgenes in the unicellular algal genus Micrasterias were developed. By obtaining axenic lines through zygospores treatment and antibiotic cocktail, and utilizing PEG-mediated protoplast transformation and particle bombardment, successful transient transgene expression was achieved. Moreover, a higher promoter activity using the 5' -flanking region of the MrACT1 gene fused with green fluorescent protein was observed in comparison with heterologous promoters.