Expanding the molecular toolbox for Zygnematophyceae - transient genetic transformation of the desmid Micrasterias radians var. evoluta

• Published in: European journal of phycology 2021-01, Vol.56 (1), p.51-60
• Main Authors: Zhou, Hong, Wilkens, Alwine, Hanelt, Dieter, von Schwartzenberg, Klaus
• Format: Article
• Language: English
• Subjects: Actin; Algae; Antibiotics; Biology; Cell differentiation; Cell size; Cytology; Cytoskeleton; Desmidiaceae; endogenous promoter; Fusion protein; Genetic transformation; Germination; Green fluorescent protein; Labeling; Life cycle; Life cycles; Micrasterias; Micrasterias radians; model organism; Particle bombardment; PEG; Polyethylene glycol; Proteins; Protoplasts; Regeneration (biological); Survival; Transformed cells; Transgenes; Vegetative cells; Zygnematophyceae; Zygospores
• ISSN: 0967-0262; 1469-4433
• DOI: 10.1080/09670262.2020.1768298

In the past, various species of the unicellular algal genus Micrasterias (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 make Micrasterias interesting, especially for cytomorphogenetic studies. However, within this genus a model organism enabling molecular research has not yet been established. Micrasterias radians var. evoluta (W.B.Turner) Krieger allows easy culturing under axenic conditions and performs its whole life cycle in vitro thus 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 the M. radians var. evoluta Actin1 gene (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 in M. radians var. evoluta.