Adventitious shoot formation from leaf explants of Rhododendron
A protocol for high frequency adventitious shoot regeneration adventitious shoot regeneration from leaf explants of Rhododendron spp. has been developed. The highest percentage of regeneration and the greatest number of shoots were obtained when leaf explants were cultured on Anderson's medium containing 4.9 μM IBA and 73.8 μM 2iP. Genotypic variation was observed for adventitious shoot regeneration potential among the seven cultivars tested. Regeneration frequencies ranged from 0 to 96%. ‘Lodestar’ had the highest rate of regeneration after 3 months of culture with 96% shoot regeneration and an average of 14 shoots per explant. Regenerated shoots were rooted in soil in about 2 months. This protocol should be useful in applying gene transfer techniques to Rhododendron improvement.
Plant regeneration from somatic tissue of Rhododendron laetum x aurigeranum
The influence of the source of plant material (greenhouse-grown plants or in vitro shoot cultures), the type of tissue explant (shoot-tip, single-node stem segment, whole leaf, leaf strip or half-leaf section) and growth regulator concentration on shoot regeneration from somatic tissue of Rhododendron laetum × aurigeranum was evaluated. No regeneration response was obtained on explants from greenhouse-grown plants. Adventitious shoots were obtained from callus produced at the basal end of shoot-tip and single-node stem segment explants derived from in vitro-grown shoots cultured on Anderson's medium supplemented with 22.8 μM IAA and 73.8 μM 2iP. The greatest percentage of adventitious shoot regeneration (77%) was induced on leaf sections cultured in the presence of 22.8 μM IAA and 147.6 μM 2iP. Plant regeneration was accomplished with minimal callus formation. This technique represents a further step toward gene manipulation of Rhododendron.
A survey of the response of Rhododendron to in vitro culture
The responses of 7 genotypes of Rhodendron to culture conditions and their establishment as shoot cultures are described. The genotypes represent a broad genetic diversity in the genus. After sterilization and an acclimation period of 3 to 12 months, all the selections were established as shoot cultures on Woody Plant Medium (WPM) supplemented with N6(-Δ2-isopenteny) adenine (2iP). Plants with strong episodic growth cycles required the longer acclimation periods. Utilizing shoots from these cultures, the response to a cytokinin series of 0 to 32 μM 2iP or BAP (6-Benzylaminopurine) was analyzed. BAP proved toxic to all but the elipidote and lepidote rhododendrons (R. mucronulatum, R. x ‘Boule de Neige’, and R. x ‘PJM’); however, even with these selections, 2iP stimulated greater shoot multiplication rates. The optimum 2iP level for shoot multiplication varied little with the genotype and levels of 4 to 16 μM generally proved optimal, depending on the specific selection. Adventitious shoot production was observed in 3 selections (R. canadense, R. x ‘Boule de Neige’ and R. x ‘PJM’), but only at 2iP levels above 8 μM. Shoot multiplication rates of 7 to 21 times were observed, depending on the selection. Using an average utilizable shoot production rate of 40 shoots per culture per 6 week subculture period, some 75,000 shoots can be generated per square meter of culture space per year. The harvested shoots (microcuttings) rooted readily out-of-culture and the resultant plants grew like seedlings.