Cryopreservation of Shoot Tips of Tetraploid Potato (Solanum tuberosumL.) Clones by Vitrification

Abstract

In vitro -grown shoot tips of five tetraploid potato ( Solanum tuberosum L.) clones were cryopreserved by vitrification. Excised shoot tips (0.5–0.7 mm) were pre-cultured on filter paper discs over half strength liquid Murashige and Skoog (MS) medium supplemented with 8.7 μ m GA 3 and different combinations of sucrose (0.3, 0.5 and 0.7 m ) plus mannitol (0, 0.2 and 0.4 m ) for 2 d under a 16 h photoperiod at 24 °C. The pre-cultured shoot tips were either successively loaded with 20 and 60% PVS 2 solutions or directly exposed to concentrated vitrification solution before physical vitrification during liquid nitrogen treatment. The vitrified shoot tips were warmed rapidly and treated with dilution mixture (MS+1.2 m sucrose) for 30 min before plating on regrowth medium. Addition of mannitol to the pre-culture medium improved survival of vitrified shoot tips. Direct dehydration of pre-cultured shoot tips with concentrated PVS 2 was detrimental to survival of vitrified shoot tips. Shoot tips pre-cultured on medium containing 0.3 m sucrose plus 0.2 m mannitol, and loaded with 20% PVS 2 for 30 min followed by 15 min incubation in 60% PVS 2 and 5 min incubation in 100% PVS 2 at 0 °C resulted in up to 54% survival after vitrification. About 50% of vitrified and warmed shoot tips formed shoots directly. Post-thaw culturing of vitrified shoot tips on medium containing an elevated level of sucrose (0.2 m ) under diffuse light for the first week enhanced the survival rate. Continuous culturing of vitrified shoot tips on high-sucrose medium induced multiple shoot formation. Copyright 1998 Annals of Botany Company