Domestication of the Nitrogen Fixing Fern-Cyanobacteria Symbiosis Azolla for the Circular Economy

Without nitrogen fertilizer Azolla filiculoides yielded 35 tons dry weight biomass/ha.year containing 24% w/w protein. A. pinnata yielded similarly. Elevated CO₂increased yields to 45 tons dry weight/ha.year with unchanged protein content. We therefore currently develop Azolla farming systems which require no nitrogen fertilizer but industrial waste for protein production to substitute soybean imports in Europe.

We demonstrated storage and dissemination control necessary for domestication and development of Azolla nurseries: A. filiculoides spore formation could be reliably induced. Spores as well as the cyanobacterial symbiont akinetes were cryopreserved. Spores were in vitrofertilized and sporelings grown.

Using Ion Torrent RNAseq of different tissues collected at four time points during the diurnal cycle allowed construction of a transcript database containing 37 k contigs of which 21 k could be annotated based on their homology to known plant genes. To validate the database we characterized genes associated with induction of sporulation. RNAseq using Illumina further quantified the response of the fern to nitrogen fertilizer at four time points during the diurnal cycle and confirmed that the fern is unresponsive to N-fertilizer compared to known crop plants.

Development of molecular breeding is impeded by the absence of genome sequence from ferns. We therefore prepared single plant derived genomic DNA from A.filiculoides that we share with the Azolla genome sequencing community and have begun sequencing using PacBioRSII towards the best possible assembly and annotation of this 750 Mb fern genome.