Cold-enhanced somatic embryogenesis in Pinus patula is mediated by calcium

The possible inter-relationship between exogenous calcium during cold pretreatment-induced somatic embryogenesis was investigated using vegetative shoot apices of mature trees of Pinus patula. Cold pretreatment of thick (0.5–1.0 mm) apical shoot sections at 2 °C for 3 days on 0.3% activated charcoal-induced white mucilaginous embryogenic tissue on initiation medium (II). The enhanced embryogenic response of cells to cold pretreatment was dependent on the Ca2+ level in the pretreatment medium (I). Addition of Ca2+ channel blockers such as La3+ (LaCl3) or ethyleneglycol-bis-B-aminoethyl ether-N,N,N,N-tetra acetic acid completely eliminated the cold-induced enhancement of somatic embryogenesis. These results suggest that embryogenic cells require minimal concentrations of Ca2+ during pretreatment for the expression of the cold-enhancement of embryogenesis in mature Pinus patula tissues.