Transformation and conjugal transfer of foreign genes into the filamentous multicellular cyanobacteria (subsection V) Fischerella and Chlorogloeopsis

Abstract

Cyanobacteria of subsection V grow as filaments with asymmetrical cell divisions that can generate a true-branching phenotype. Members of the genera Fischerella and Chlorogloeopsis furthermore differentiate akinetes (spore-like resting stages), heterocysts (specialized in nitrogen fixation) and hormogonia (cell aggregates with gliding motility for colonization and dispersal). Genetic approaches to studying the complex morphology and differentiations of these prokaryotes require transformation techniques. For Fischerella and Chlorogloeopsis reliable protocols for introducing foreign genes are lacking. Here, we explored conjugation, electroporation, and biolistic DNA transfer methods in Fischerella and Chlorogloeopsis, using the cyanobacterial replicon pRL25C as a marker. We successfully transformed Fischerella muscicola PCC 7414 and Chlorogloeopsis fritschii PCC 6912 and were able to express the GFP reporter protein under two different promoters: the nitrogen regulated (p) glnA and the strong E. coli hybrid (p) trc. For Fischerella all methods worked, for Chlorogloeopsis electroporation was unsuccessful. For both strains conjugation delivered the most reproducible results, whereby partial removal of the exopolysaccharide sheath by salt washing was a critical step.