Cryopreservation of embryogenic cultures of Scots pine

The aim of the study was to develop an effective cryopreservation method fo r Scots pine (Pinus sylvestris L.) embryogenic cultures. Altogether nine ce ll lines derived from three mother trees were cryopreserved after cold hard ening using dimethylsulfoxide or two different mixtures of polyethyleneglyc ol 6000, glucose and dimethylsulfoxide as cryoprotectants. Seventy-eight pe rcent of the cell lines remained viable after cryostorage, the best cryopro tectant treatment being 10% polyethyleneglycol 6000, 10% glucose, and 10% d imethylsulfoxide in water. This treatment resulted in significantly better regrowth of the embryogenic cultures than with the other cryoprotectants or with the controls. According to microscopical observations, the cells that retained their viability and regrowth ability after cryopreservation were the embryonal head cells, as well as some elliptic suspensor cells close to the embryonal head cell area. When proliferation growth of the frozen cult ures had started, their morphological appearance was the same as the non-fr ozen cultures. In addition, the RAPD assays suggested that the cryostorage treatment used here preserved the genetic fidelity of the Scots pine embryo genic cultures.