PROLIFERATION AND COLD HARDENING OF IN VITRO GROWN APPLE SHOOT TIPS

Abstract

Shoot tips of apple cultivar 'Rubin' were successfully established in vitro. Five proliferation MS (Murashige and Skoog) media containing different concentrations of BAP (0.25, 0.5, 1, 2 mg L -1 ) or a combination of 1 mg L -1 BAP and 0.1 mg L -1 IBA were tested. The highest proliferation rate was obtained on MS medium with 1 mg L -1 BAP, which produced 4.2 ±0.3 shoots/explant after one month of cultivation. In vitro plants of cv. 'Rubin' were cultivated for 4 weeks at low temperature (4°C) and short photoperiod (8/16-h - light/dark). The assessment of frost resistance of cold hardened and non-hardened in vitro apple plants was done by freezing tests. In vitro plants were exposed to the controlled cooling rate 4°C/h. The lethal temperature (LT) was -13.0°C for cold hardened and -6.5°C for non-hardened in vitro plants. Cold hardened in vitro cultures were used for subsequent cryopreservation. Cryopreservation was made by encapsulation-dehydration vitrification method. The survival and regeneration of cv. 'Rubin' was 78% after thawing in 40°C water.