High-efficiency transfection ofAcanthamoeba castellaniiusing a cationic polymer

Abstract

The free-living amoebaAcanthamoeba castellaniiis an ecologically, clinically, and evolutionarily important microorganisms.A. castellaniiamoebae are directly pathogenic to humans, and serve as reservoirs for bacterial pathogens (e.g.,Legionella pneumophila), but also regulate the proliferation of other microorganisms in the soil. Despite their importance, no reliable genetic system has been developed, hampering the use ofA. castellaniiand related species as model organisms. TransfectingA. castellaniiwith plasmids is possible with commercial kits, but is expensive, inefficient, and vulnerable to product discontinuation. In this contribution, we present a method for efficient transfection ofA. castellaniiwith readily available and inexpensive polyethylenimines. We systematically explore the parameters of the method, obtaining up to 100-fold higher efficiency than currently used reagents. The method presented here provides a robust step towards a full genetic toolbox forA. castellanii, hence expanding its use as a model organism.