Cryopreservation of immature and mature seeds of Brazilian orchids of the genus Cattleya

Abstract

The aim of this study was to evaluate the efficiency of cryopreservation by the vitrification technique as a method of long-term conservation of immature seeds of the Cattleya harrisoniana × Cattleya walkeriana hybrid and of Cattleya tigrina with high water contents (49.35% and 65.60%, respectively) and mature seeds of Cattleya guttata with a low water content (6.15%). For each species or hybrid, the experiment was conducted in a completely randomized design with five treatments, namely, direct seed germination in vitro without PVS2 and without liquid nitrogen; PVS2 at 0 °C for 60 min without liquid nitrogen; PVS2 at 0 °C for 30 min or 1 h, followed by immersion in liquid nitrogen; and direct immersion of seeds in liquid nitrogen without PVS2. Five replicates were used. Each replicate was composed of a cryotube containing 15 mg of seeds. The following variables were evaluated after cryopreservation: seed viability, seed germination, seedling growth in vitro and seedling growth after acclimatization in a greenhouse. Plant genetic stability was examined by flow cytometry after in vitro culture. Anatomical analyses were carried out on the plants after acclimatization. Cryopreservation by vitrification was shown to be an effective and necessary method for the conservation of immature seeds of the C. harrisoniana × C. walkeriana hybrid and of C. tigrina with high water contents. For mature seeds of C. guttata with a low water content, direct immersion in liquid nitrogen without PVS2 was effective, and the use of vitrification was not required. These results demonstrate the effect of seed water content on cryopreservation. This unprecedented work examines the cryopreservation of the C. harrisoniana × C. walkeriana hybrid, C. guttata and C. tigrina from pollination to the production of acclimatized plants.