In vitro culture and medium-term conservation of the rare wild species Gladiolus imbricatus

Abstract

Gladiolus imbricatus, a rare species spread in the hill region of Europe, is resistant to abiotic and biotic stress being one of the most cold-tolerant in the genus. Moreover it contains high vitamin C and minerals in the leaves and the flowers are considered as edible. The aim of our study was to develop in vitro technologies for micropropagation, multiplication, corm development, somatic embryogenesis and medium-term storage of this endangered species, Red listed in South-Eastern and Central European countries. Initiation of in vitro cultures was started from wounded seeds or excised mature embryos. Micropropagation and multiplication was achieved on Murashige Skoog (MS) solid medium with 30 g L^-1 sucrose and 1 mg L^-1 N-6-benzyl adenine (BA). The callus developed from wounded seeds on a richer MS medium proved to be embryogenic. Increased concentrations of sucrose promote corm development as also shown for other genotypes of cultivated gladioli. Further on a two-step culture protocol, on solid and liquid shaken MS based media, each for 6 months was done to evaluate the effects of acetic acid and ancymidol on corm development. Acetic acid + sucrose 9%, alone or in combination with ancymidol stimulated corm formation and yield. For medium term conservation maintenance of the in vitro cultures in low temperature in the dark proved to be the best in reducing the growth rate of the shoots after 3, 6 and 12 months. The recovery of plant growth was evaluated after 12 months, by the transfer to normal growth conditions. 25% of the plants were recovered after one year storage. Besides the importance for biotechnology, the in vitro techniques described here might be used, after molecular analysis of genetic stability, for the restoration of natural populations into the habitats where this species became extinct.

Keywords: Acetic acid, ancymidol, corm development, somatic embryogenesis