Immature and mature nonstratified seeds of white ash (Fraxinus americana L.) were dissected transversely and 2/3 of each seed was placed onto agar-solidified Murashige and Skoog medium. Adventitious buds, shoots, and somatic embryos formed on callus, cotyledons, and hypocotyls of the resulting seedlings. Shoot organogenesis was induced on explants cultured on medium with 10 txM thidiazuron but not on explants on media with benzyladenine (BA) or isopentenyladenine. Not all seed sources were equally capable of shoot organogenesis and embryogenesis. Atypical of adventitious regeneration of other woody plants, mature seed explants of white ash were more organogenic with shoots that elongated better than explants from immature seeds. Somatic embryogenesis was observed in cultures where mature seeds were first cultured for 4 weeks on a medium containing 10 I~M adenine 2,4-dichlorophenoxyacetic acid in combination with 0.1 and 1.0 IxM thidiazuron, followed by transfer to a medium containing 0.05 ~M 6-benzyladenine and 0.5 IxM naphthaleneacetic acid. Adventitious shoots and epicotyls from both seedlings and germinated somatic embryos were rooted under intermittent mist and acclimatized to the greenhouse.
Axillary shoots from three selected white ash (Fraxinus americana L.) clones were harvested from in vitro shoot cultures. Roots were initiated by pulsing excised shoots for eight days in the dark in MS medium supplemented with 2% sucrose, 0.7% agar, 5 k~M NAA, and 1/xM IBA. Pulsed shoots were transferred to a root elongation medium consisting of 25% MS macrosalts, full-strength microsalts and organics, 1% sucrose, 0.7% agar and no auxins. When roots were visible (6-10 days after transfer to root elongation medium), microplants were transferred to vessels containing the same minimal medium and tall fescue (Festuca elatior var. arundinacea (Schreb.) Wimm.) leaf extracts, leaf leachates, or soil leachates from plant boxes with and without tall fescue sod. After four weeks in vitro, primary adventitious and secondary root growth was reduced by extracts obtained from 5 and 10 g ground leaves per 100 ml of medium. Leachates obtained from 5 g soaked leaves per 100 ml of medium stimulated primary root growth. Soil leachates from bare soil also stimulated primary root growth. Variation was observed among the clones for root growth when plantlets were grown in extracts or leachates from tall fescue.
On 10 Apr. 1997, the smaller branch on forked Fraxinus americana L. (white ash) trees in an 8-year-old clonal plantation were removed, cut into 25-cm-long stem sections, and placed horizontally in perlite under seven different forcing regimes. Sprouts from latent buds were excised 67 and 99 days later and trimmed to retain the apical pair of leaves and terminal bud. The basal 2 cm of each cuttings was set for 30 to 60 min in one of six aqueous dilutions made from a stock solution of 1% IBA and 0.5% NAA (Dip `n Grow) before placing in a 1 perlite: 1 vermiculite medium. Eight weeks after treating, cuttings data were collected. Most cuttings treated with 3200 mg/L IBA plus 1600 mg/L NAA quickly died. Survival (85%) and rooting percentage (86%) were similar for the remaining auxin-treated cuttings and controls with water only. Cuttings treated with 1600 mg/L IBA plus 800 mg/L NAA produced the most adventitious roots (6.2 roots), the longest adventitious root (22 cm), and longest combined sum of all adventitious roots (51 cm); however, these rooted cuttings died when transplanted to soil. Cuttings treated with 100 mg/L IBA plus 50 mg/L NAA or 400 mg/L IBA plus 200 mg/L NAA had more adventitious roots (3.2 roots) and total length of adventitious roots (370 mm/cutting) than cuttings treated with 40 mg/L IBA plus 20 mg/L NAA or controls with water only (2.2 roots, 220 mm long). Results indicate softwood cuttings forced on stem segments of adult white ash readily root under mist following a brief soak in a 400 mg/L IBA plus 200 mg/L NAA solution.
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