Establishment of a carrot cell plant growth regulators-free system to produce a low-cost vac-cine candidate.

Parkinson´s disease is characterized by an abnormal accumulation of alpha synuclein (α-syn) in dif-ferent regions of the central nervous system. At present, pharmacological treatment is effective, but it is a palliative condition. Immunotherapy is considered an alternative to treat Parkinson´s disease, and plants are a convenient alternative platform for biopharmaceutical production. When compared to other systems, plants are particularly attractive because they offer cost-effectiveness, large-scale production, and enhanced safety. Therefore, this study aimed to establish a carrot cell suspension culture for the production of cLTB-Syn, a vaccine candidate against Parkinson´s disease. The con-venience of MS medium optimization was demonstrated. Transgenic callus cultures were maintained and adapted on solid MSU9 medium without phytohormones, followed by growth kinetics in suspen-sion cultures. The maximum biomass yield was 15.8 ± 0.35 g/L DW at 14 days of culture, with a growth rate of µ= 0.1034/d and td=6.7 days. The cLTB-Syn protein production reached a maximum value of 2.62 ± 0.03 µg/g DW, representing a 1.6-fold increase over the initial culture time. Finally, the presence of the transgene was confirmed by PCR, and the integrity of cLTB-Syn protein was deter-mined by dot blot assays. This study presents evidence of a promising system for a toxin free bio-pharmaceutical production, which has the potential to be scaled up for large manufacturing, at a low cost.