Improvements to Solanum tuberosum cv. ‘Desireé’ cell suspension culture and Nicotiana tabacum cv. ‘Petit Havana’ shoot induction.

Since the 1980s, progress in biotechnology has harnessed the incredible potential of plant biology. Plants have been engineered to be pest and herbicide resistant, enhance stress tolerance, and produce pharmaceutical proteins. Despite these feats of genetic engineering, plant tissue culture remains a limiting factor for future research. Unfortunately, the pressure to quickly innovate and produce novel products has pushed aside research to optimize tissue culture and exploring underlying molecular mechanisms for improved culture conditions. Efficient tissue culture methods are limited to a few plant taxa and are sometimes described in an oversimplified manner under the assumption they are facile to replicate. As a result, nuances of methodology, that are difficult to quantify, remain undescribed; introducing limitations in reproducibility across differing laboratory environments. Two such examples of overlooked plant tissue culture tools are potato cell suspension culture and rose bengal [4,5,6,7-tetrachloro-2',4',5',7'-tetraiodofluorescein (RB)]. Potato cell suspension cultures, although prevalent in literature, remain relatively uncharacterized. RB is a red, xanthene dye that produces reactive oxygen species (ROS) under light conditions. RB, although supported by literature as a promising tool in plant ROS research, is distinctly unexplored in plant tissue culture. To address this dearth in scientific literature, potato cell suspension culture was optimized and the effects of RB on tobacco plant regeneration were characterized. Potato cell suspension optimization established that Solanum tuberosum cv. ‘Desireé’ friable callus induction is dependent on auxin, internodal callus is more productive, producing between 2.9- and 4.8-fold more biomass, on solid media than leaf callus, leaf callus sourced cell suspension culture are about 1.2-fold more productive than internodal counterparts and that the optimized culture produced by this work is highly replicable. The effect of RB on tobacco shoot regeneration was evaluated and the results demonstrated shoot induction was enhanced between 1.7- and 2.2-fold at concentrations of 10-30 mg/L, inhibited at concentrations of 80-100 mg/L and decreased between 3.9- and 9.4-fold at sub-lethal concentrations of 50-70 mg/L. Overall, statistically significant improvements to and valuable insights on the replicability of potato cell suspension as well as the ability to utilize RB as a tool in plant tissue culture were elucidated.