Establishment of an ex vivo laticifer cell suspension culture from Taraxacum brevicorniculatum as a production system for cis-isoprene

Laticifers are highly specialized plant cells that produce latex enriched with secondary metabolites. The articulated laticifers of Taraxacum brevicorniculatum synthesize natural rubber, an industrially-valuable composite biopolymer comprising >95% high-molecular-weight (HMW) poly(cis-1,4-isoprene). Here we present a proof-of-concept approach for the cultivation of cell suspension cultures exclusively containing laticifers. We transformed T. brevicorniculatum plants with a plasmid conferring laticifer-specific hygromycin resistance. Transgenic callus tissue was used to induce a cell suspension culture under antibiotic selection to favor laticifer growth. The cultured cells appeared laticiferous in terms of morphology and expressed laticifer-specific genes. Confocal laser scanning microscopy revealed intracellular lipid accumulation in vesicle-like structures. Nuclear magnetic resonance and diffusion ordered spectroscopy indicated the presence of mid-length poly(cis-1,4 -isoprene) chains but no high HMW natural rubber in the cells. Precursor feeding with mevalonolactone increased the accumulation of poly(cis-1,4-isoprene) by 17-fold, reaching a concentration of 2.7 mg/g dry weight. Our approach could lead to the development of a production platform for the efficient conversion of isopentenyl diphosphate into poly(cis-1,4-isoprene) in an optimized cell suspension culture system. The apparent absence of HMW natural rubber is discussed in terms of our current knowledge of rubber biosynthesis