Cryopreservation of in vitro grown shoot tips of two interspecific Prunus rootstocks

Abstract

Shoot tips of two in vitro grown interspecific Prunus rootstocks, Fereley-Jaspi (R) and Ferlenain-Plumina (R), were successfully cryopreserved using a two step freezing method. Excised shoot tips from hardened plantlets were precultured for 24 h on solid medium (modified half strength Murashige and Skoog medium (MS)) supplemented with 5% dimethylsulfoxide and 2% proline. They were transferred to 1.2-ml cryotubes and treated with 0.5 ml of modified PVS2 cryoprotective solution for 20–40 min. Cryotubes were then cooled at 1°C/min to −40°C before being immersed in liquid nitrogen (LN2). After 1 min warming at 40°C, shoot tips were rinsed with MS medium containing 1.2 M sucrose and then plated. First regrowth occurred after 5 days and was evaluated 21 days after plating. The average rate of shoot formation was 69% and 74% for Fereley and Ferlenain respectively, compared with 86% for controls of both hybrids. The roles of several factors, especially hardening procedures and meristem size, in the improvement of survival rates, were discussed.