Optimization of a working cryopreservation protocol for Pinus patula embryogenic tissue

Summary The protocol currently used to cryopreserve Pinus patula embryogenic tissue was investigated in an attempt to improve and optimize the recovery of tissue. This investigation describes two aspects that influence tissue recovery after cryopreservation: (i) the effects of precooling tissue prior to immersion into liquid nitrogen; and (ii) whether the choice of supports onto which the recovered tissue is suspended improved the recovery rate. Results indicated that precooling tissue to −70°C prior to immersion into liquid nitrogen was superior to precooling to −30°C or direct immersion in liquid nitrogen (−196°C). Tissue recovery improved when polyester grids were used as supports, and was slowest on filter paper supports.