Dormant meristems of fascicles explanted from 10-year-old, field-grown trees of Pinus brutia Ten. were cultured in vitro. Browning of cultured fascicles was reduced by including 150 mg l(-1) sodium diethyldithiocarbamate (SDD) in a cytokinin-containing medium. The stage of development of fascicles when placed in culture affected both shoot-bud production and the degree of browning. Only fascicles at an advanced stage of development had a high rate of shoot-bud production. Fascicles cultured for 6 weeks in initiation medium containing 150 mg l(-1) SDD and then for 4 weeks in initiation medium containing 1 mg l(-1) insoluble polyvinylpolypyrrolidone showed the highest rate (23%) of shoot-bud development and the lowest rate (15%) of browning. Elongation of activated shoot buds was considerably enhanced by reducing the concentration of cytokinin in the culture medium. Only shoots >/= 20 mm in height were capable of producing further crops of buds and shoots. When treated with a combination of auxin and cytokinin, only 16% of the elongated shoots produced roots.

SUMMARY The effect of auxins and a cytokinin on induction of roots in cultured axillary shoots of Pinus brutia Ten, has been tested. Auxin was crucial for root initiation and the rooting response varied according to the type and concentration of auxin applied. Both auxin and cytokinin and the interactions between them affected the quantity and quality of the induced roots. Aerated non‐sterile tap water was an effective rooting medium, comparable to agar. After planting out into soil, some of the influences of auxin and cytokinin could still be seen after six months. However, roots developed normally in the soil, displaying dichotomous lateral branches. The results draw attention to the need for care in the choice and application of the medium for the initial induction of roots. Results of greenhouse trials indicated that the most vigorous plants were obtained via the axillary shoots.

Axillary shoot production was achieved in 6 weeks using excised shoot explants of Pinusbrutia Ten. on a modified Schenk and Hildebrandt medium containing cytokinin. Primary shoots arose from existing axillary buds and secondary buds arose from bases of the primary shoots. Their production could be increased by regulating the cytokinin level and by surgical removal of apical buds from the cultured explants. However, the best performance was achieved with a low level of 6-benzylaminopurine (3 × 10−6 M) or a mixture of 6-benzylaminopurine and kinetin (10−6 M of each). Subsequent transfer to a cytokinin-free medium resulted on average in the production of 43 shoots per cultured explant and up to 67 shoots per clone within 12 weeks. When the primary shoots, which had already produced one crop of secondary shoots, were maintained under conditions favourable for shoot production, a doubling in number was obtained within 4–6 weeks. To encourage further elongation, newly formed shoots were incubated for 2 weeks on a cytokinin-free medium to which 1% activated charcoal was added. The time taken with this method was much shorter than with other published methods and is, therefore, likely to be important for the vegetative propagation and multiplication of selected seedlings of this species.

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In vitro adventitious shoot formation from embryonic and cotyledonary tissues ofPinus brutia Ten.Abstract. Adventitious buds were induced when isolated whole embryos, and excised cotyledons from treated seeds of Calabrian pine (Pinus brutia Ten.) were cultured on a cytokinin-supplemented medium. The adventitious buds formed dkectly from the cotyledons. The highest number of bud primordia were formed, with both embryos and excised cotyledons, after 6 weeks on a BAP -supplemented Schenk and Hildebrandt medium under a 16 h photoperiod. The buds, when separated and maintained individually on a full-strength medium without growth regulators, developed into well-formed shoots within 4 weeks. The average number of harvested shoots obtained (> 1 cm in height) per seed over 24 weeks was 55; however, a maximum number of 152 shoots was obtained from one individual over the same period. The shoot forming capacity of the meristematic tissue was not lost after seven harvests.