Cultural factors influencing adventitious shoot and plantlet formation from slash pine cotyledons

Cultural factors affecting in vitro shoot and subsequent plantlet formation of slash pine (Pinus elliotti Engelm.) cotyledons were investigated. Basal media composition, N⁶-benzylaminopurine (BAP) concentration and exposure time significantly influenced bud induction in cotyledons cultured under a continuous photoperiod of 35–40 mol m^–2 s^–1 at 24 ± 1 °C. The largest number of adventitious shoots was obtained after 28 days exposure to 66 M BAP-supplemented modified Gresshoff and Doy 1 (GD1) medium. Relatively high frequencies of large shoots were obtained after a 14-day exposure to 22 M BAP-supplemented Brown and Lawrence (BL) or 66 M BAP-supplemented GD1. Adventitious shoots derived from 21- or 28-day exposures to BAP developed more slowly and were smaller in size than those derived from a 14-day exposure to the cytokinin. Shoot differentiation and subsequent growth were also influenced by basal media, media concentration, and presence of activated charcoal in the medium. The percentage of cotyledons forming shoots was highest on half-strength GD1 medium containing activated charcoal. Rooting was achieved in vitro under a continuous photoperiod of 60–70 mol M^–2 S^–1. Roots were formed when excised shoots were planted on GD 1/2 medium supplemented with 2.68 M 1^–1 a-napthaleneacetic acid (NAA) with or without BAP for 14 days. The proposed technique of slash pine propagation using cotyledon explants can produce up to 100 seedlings per embryo.