THE ORIGIN AND DEVELOPMENT OF SOMATIC EMBRYOS FOLLOWING CRYOPRESERVATION OF AN EMBRYOGENIC SUSPENSION-CULTURE OF PICEA-SITCHENSIS

The development of somatic embryos in an embryogenic suspension cultur e of Picea sitchensis was followed every day for two weeks after thawi ng from liquid nitrogen (LN(2)). Only a few cells, primarily located a l the periphery of the embryonic region of the embryos, survived cryop reservation in LN(2). Surviving cells were classified into two groups: embryogenic cells (EC) and non-embryogenic cells (NEC), based on thei r morphology and embryogenic competence. The dense cytoplasmic EC unde rwent organized growth and differentiation with first divisions occurr ing after 24 h, and embryo formation 6-8 days after thawing from LN(2) . No evidence of asymmetrical divisions or free-nuclear stages was fou nd during somatic embryo formation. NEC had less dense cytoplasm with numerous small vacuoles. One to five days after thawing the NEC became progressively more vacuolated and elongated. Histological examination revealed no mitotic activity in NEC, and six days after thawing NECs were seen as single cells or unorganized cell aggregates. Two weeks af ter thawing the appearance of the cryopreserved cultures was comparabl e to that of the untreated cultures.