A protocol is described for plantlet regeneration using embryonic explants of Juniperus cedrus Webb & Berth.An average of 6 adventitious buds were induced on whole excised embryos cultured for 15 days on Quoirin and LePoivre (QP) half-strength medium supplemented with 5 laM N6-benzyladenine. For bud development, explants were transferred to phytohormone-free 1/2 QP medium. For shoot elongation, explants were cultured on 1/2 QP with 0.05% activated charcoal and 2% sucrose. Adventitious shoots were rooted successfully in peat-vermiculiteperlite (1:1:1) moistened with 1/4 QP containing 1% sucrose and 5 laM a-napthaleneacetic acid, pH 5.0. Axillary shoots elongated spontaneously in culture from leaf axils.

A protocol is described for plantlet formation in juvenile tissues of Pinuscanariensis Sweet ex K. Spreng. (Canary Island pine). Adventitious buds were induced on 3-day-old cotyledonary explants cultured on Bornman's MCM medium supplemented with cytokinin. The concentration of benzylaminopurine, the use of other cytokinins alone or in combination with benzylaminopurine, and the concentration of mineral salts strongly affected the bud forming capacity of the cotyledonary explants. Also, the age of the explants significantly influenced the frequency of adventitious bud formation. Bud development was enhanced by the elimination of phytohormones, a reduction of mineral salts and sucrose, and the inclusion of activated charcoal in the medium. The conditions used during the induction phase strongly affected the ability of the induced buds to develop into vigorous rootable shoots. Vitrification problems were eliminated by transferring the shoots to the elongation medium solidified with Gelrite, and shoot remultiplication was enhanced by removing the apical bud. Maximum rooting was obtained by pulsing shoots with a high concentration of indolebutyric acid and by using peat–vermiculite or peat–vermiculite–perlite as substrates. Roots developed within 6–8 weeks, and the regenerated plantlets were transferred to soil under nonsterile conditions, where further development occurred.

A method is described for using liquid pulsing as an alternative to the conventional induction protocol for Pinus canariensis. Using Day 0 and Day 3 explants, the best exposure time was 8 h and 4 h respectively, in a non-buffered 100 ~M N6-benzyladenine solution, followed by culture on half-strength Bornman's medium containing 3% sucrose and 0.8% Difco Bacto R agar. With this procedure, 97% of the cotyledonary explants produced about 14 buds/explant. These results were comparable to a 14-day induction period on full-strength Bornman's medium containing 10 poM N6-benzyladenine.

Three-day-old cotyledonary explants of Pinus canariensis were subjected to 30 induction treatments using half-strength Bornman's medium containing various combinations of N 6-benzyladenine, zeatin, kinetin and 2-isopentenyl-adenine. The highest numbers of buds were obtained with 10 ~tM 6-benzyladenine, but both kinetin and zeatin influenced shoot elongation. Shoots were maintained on half-strength Schenk and Hildebrandt medium with 2% sucrose and 0.05% activated charcoal. For rooting, shoots were pulsed for 4 h in a 100 gM indole-3-butyric acid aqueous solution (pH 4.2-4.5), and planted in peat:vermiculite:perlite (1:1:1). After 8 weeks, the numbers of rooted shoots were similar for most treatments. Therefore, the bud induction treatments did not significantly influence rooting of adventitious shoots of Canary Island pine.

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