Establishment of methods of disinfection and callogenesis from needle explants of Abies koreana

Korean fir (Abies koreana), one of the important native conifers belonging to Pinacea and popular as Christmas trees, is a Korean endemic species distributed in the southern regions of Korea. The population of A. koreana, has been declining, probably due to the climate change, and now it is one of the rare plants that needs to be protected and conserved. The needle explants of Abies koreana were collected from lateral buds of one-year-old shoots after the winter dormancy period in May at about one month after needles sprouted out from the leaf buds. The needle explants were first dipped for 5 min in 400 mL tap water contained in a 500-mL container with 1 mL Tween 20, and then were rinsed for 3 h in running tap water before being used for different disinfection treatments. The treatments used for disinfection were combinations of different exposures to 0.5 or 1.0% (v/v) NaClO for 5 min, 70% of EtOH for 1 min and rinsing with sterilized water, and distilled water was used as the control. The explants were cultured in the MS medium in a culture room with day/night temperatures of 24/18°C and relative humidity (RH) of 80% in the dark. The treatment using 0.5% NaClO for 5 min plus 70% EtOH for 1 min followed by 3 rinses with sterilized water gave the lowest ratios of contamination (0.0%) and browning (29.2%) of the explants after 2 weeks of culture. Then the feasibility of callus induction from the disinfected needle explants on three media supplemented with plant growth regulators (PGRs) was studied. The disinfected needle explants were cultured in one of the media, each supplemented with PGR combinations, in a culture room with the same environmental conditions as shown above. The callus induction was affected significantly by the medium and PGRs. The needle explants in the MS or DCR medium supplemented with 1.0 mg L^‑1 2,4-dichlorophenoxyacetic acid (2,4-D), 2.0 mg L^‑1 1-naphthaleneacetic acid (NAA) and 1.0 mg L^‑1 kinetin (KT) gave the greatest callus induction (56.7%) and the lowest callus browning (0.0%) ratios after 4 weeks of culture. The greatest fresh weight of callus (87.0 mg) after 8 weeks of culture was obtained in the MS medium supplemented with 2.0 mg L^‑1 NAA, 0.5 mg L^‑1 KT and 2.0 mg L^‑1 benzyl-aminopurine (6-BA). These results may lay a foundation for the micropropagation and biotechnological applications of the endangered species Abies koreana.