Microcallus growth from maize protoplasts

Maize (Zea mays L.) protoplasts obtained from Type I and Type II calli from several genotypes were shown to be capable of synthesizing cell walls and forming small clusters of cells. The medium used also supported cluster formation from protoplasts obtained from root tips. The effects of various additions to the medium (such as casein hydrolysate, coconut water, amino acids, sugars, phytohormones, nitrate, calcium, and dimethylsulfoxide as well as pH variations on cellcluster formation were determined. The method of culture (protoplasts plated in agarose or supported in alginate beads in liquid medium) as well as several components of the medium were found to be critical for microcallus formation. Protoplasts obtained from embryogenic Type I callus and cultured in the medium of C. Nitsch and J.P. Nitsch (1967, Planta 72, 355–370) modified by various additions (NN 67-mod medium) were affected most by various sugars, casein hydrolysate, coconut water, and a combination of the auxins napthalene-1-acetic acid (2 mg/l) and 2,4-dichlorophenoxyacetic acid (0.1 mg/l), and the cytokinin N⁶-benzylaminopurine (0.5 mg/l). Cluster size in the agarose culture system was from 0.1 to 0.5 mm diameter and in the alginate culture system, up to 2.0 mm diameter.