Phenology of Brainea insignis was investigated at Puli, Nantou Shihkung area in Taiwan (23°59’ N, 121°00’E；Alt. 790 m). Phenology of Brainea insignis could be classified into the following phases: 1) main frond emergence in April, 2) frond expansion, spore maturation, and spore release in May, 3) slow growth from June to November, and 4) standstill growth from December to March, when the plants exhibit no obvious growth. In the wild, plants of Brainea insignis could form trophosporophyll when stem diameter reached 7 cm. More trophosporophylls were observed in plants with stem diameter wider than 10 cm. New frond number and trophosporophyll number increased with increasing stem diameter.
Spore solution of Brainea insignis were dripped in plastic boxes with soilless mix, each drop contains about 300 spores. The effects of temperature (20, 25, and 30 ℃) and nutrient solution concentration on gametophyte development and sporophyte formation were studied. A temperature of 25 ℃ was found to be the optimum temperature for growth of gametophyte and sporophyte formation. After sowing for one week under 25 ℃ conditions, spores germinated and developed to two dimensional prothalli. After sowing for three weeks, apical meristem formed in the gametophyte. Antheridia and archegonia were observed in gametophytes after sowing for five weeks. First young sporophyte was recorded after sowing for ten weeks. A total of 178 sporophytes per box were recorded after sowing for 14 weeks. Different concentrations of Johnson’s solution (0%, 25%, 50%, and 100%) were incorporated to the sowing mix before sowing, only treatment with 100% Johnson’s solution formed sporophytes after sowing for 11 weeks. Spores germinated and formed gametophytes but did not form sporophyte in other treatments and were dead after sowing for 11 weeks.
Young sporophytes of Brainea insignis with 3 to 4 unfolded fronds were grown under various day/night temperature regimes (15/13, 20/15, 25/20, 30/25, and 35/30 ℃) for 198 days. Sporophytes grew faster under 30/25 and 25/20 ℃, and started to form trophosporophylls with 22 fronds. In contrast, growth of sporophytes was slow under 20/15 and 15/13 ℃, where the fronds turned red and no trophosporophyll formed. No sporophyte was survived under 35/30 ℃ after 120 days. Plants under 25/20 ℃ produced the most trophosporophylls. Moreover, after weekly fertigation of distilled water or 25%, 50%, or 100% Johnson’s solution for ten weeks, sporophytes fertigated with 50% Johnson’s solution resulted in most frond number, stem diameter and trophosporophylls, but sporophytes given distilled water turned yellow. Thereby, young sporophytes of Brainea insignis should be grown under day/night temperature 25/20 ℃, and fertigated with 50% Johnson’s solution weekly, which resulted in better growth.
Young sporophytes of Brainea insignis were fertigated with Johnson’s solution containing various nitrogen (N) concentrations. Plants at 12 mM N produced the maximum frond length and width, stem diameter, shoot and root dry weights, and trophosporophylls. Reduced growth was recorded in plants at 16 mM N concentrations. No trophosporophyll was seen in plants at 0 or 4 mM N treatments, while 12.5 % of plants formed sori at 8 mM N. Plants at 12 mM N had highest frond appearance rate and started to expand trophosporophylls after eight weeks of treatment. Half plants formed trophosporophyll at 12 or 16 mM N treatments. The 12 mM N treated plants had more pinnule with sori, while the 16 mM N treated plants exhibited wrinkled trophosporophylls.