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 culture of Picea sitchensis was followed every day for two weeks after thawing from liquid nitrogen (LN(2)). Only a few cells, primarily located al the periphery of the embryonic region of the embryos, survived cryopreservation in LN(2). Surviving cells were classified into two groups: embryogenic cells (EC) and non-embryogenic cells (NEC), based on their morphology and embryogenic competence. The dense cytoplasmic EC underwent organized growth and differentiation with first divisions occurring after 24 h, and embryo formation 6-8 days after thawing from LN(2). No evidence of asymmetrical divisions or free-nuclear stages was found 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 after thawing the appearance of the cryopreserved cultures was comparable to that of the untreated cultures.