Cryopreservation strategies for Cyathea australis (R. BR.) domin.

The influence of liquid nitrogen (LN) on the germination of C. australis spores and survival of gametophytes at various stages of development was investigated. Exposure to LN did not change the viability of mature spores (80 percent) but stimulated the germination of immature spores from 1.9 percent to 41 percent. Disinfection before cryopreservation contributed to loss of spore survival. However, some germination capacity was regained (48 percent) if the sterilized spores were enclosed in alginate capsules and subsequently exposed to osmotic desiccation and 5-hour air drying. Development of gametophytes derived from frozen and non-frozen spores was similar. Preculture factors (the period, type and abscisic acid treatment) affected gametophyte viability and growth. A two week preculture on agar significantly increased survival compared to preculture in a liquid medium. Addition of abscisic acid (ABA) to solid or liquid media stimulated explant survival. Highest viability (85 percent) of frozen-thawed gametophytes was achieved by a 2-week preculture in agar with 0.25 M sucrose and 10 muM ABA. Gametophytes developed directly from spores grew and multiplied in vitro at a uniform rate. Young, intensively growing gametophytes and large, proliferating ones survived better (73-80 percent) following cryoexposure than mature, non proliferating gametophytes (50 percent). Less than one quarter of the explant surface was alive in 60-80 percent of the gametophytes that survived cryoexposure.