Medium and calcium nitrate influence on shoot regeneration from leaf explants of guayule

A comparison of macro and micro nutrient levels in MS, DKW, and WPM media revealed significant differences in the calcium levels. Under controlled conditions, guayule plants in vitro incorporated calcium into plant tissues in proportion to the amount available. Moreover, the addition of calcium nitrate tetrahydrate at moderate levels into MS medium significantly improved the emergence of single, elongated true shoots, suggesting a requirement for calcium in guayule tissue culture. We have further explored selection agent (antibiotic) concentration for the identification of transgenic plants using a spectinomycin resistance gene (aadA). Based on the results, we have improved tissue culture protocols for guayule including spectinomycin as the selection agent.