Investigating Prevalence, Induction, and Fertility of Polyploid Rhododendron L. and the Development of Protocols for Vegetative Propagation

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Studies were conducted to determine the ploidy levels of specific Rhododendron taxa, to develop a simple and effective ex-vitro method for inducing polyploidy in Rhododendron seedlings, and to evaluate the effect of increased ploidy level on pollen fertility. A diverse collection of species, hybrids, and cultivars in the Hymenanthes (elepidote rhododendrons), Rhododendron (lepidote rhododendrons), Pentanthera (deciduous azaleas), and Tsutsusi (evergreen azaleas) subgenera were surveyed to determine ploidy level and relative genome size using flow cytometry. In instances where ploidy levels were inconsistent with past literature, chromosome counts were made on young root tips to substantiate findings. Mean 2C holoploid genome sizes varied as a function of subgenus and ploidy level. Relative genome sizes (2C) within ploidy level, for a given subgenus, had a narrow range providing clear distinction among ploidy levels. Mean 1Cx monoploid genome size was conserved across ploidy levels within a subgenus. Polyploidy was found to be common in the genus Rhododendron and considerably more prevalent in the subgenus Pentanthera than previously known. Particularly noteworthy were the findings that R. occidentale includes both diploid and tetraploid individuals and that R. atlanticum and R. austrinum are predominantly tetraploid species. Induction studies were then completed with the goal of obtaining artificial tetraploids from diploid x diploid hybridizations. The effectiveness of using repeated treatments of an oryzalin suspension in a warm agar solution applied directly to apical shoots of Rhododendron seedlings to induce polyploidy was tested. Apical meristems of hybrid seedlings were subjected to 1, 2, 3, or 4 applications of oryzalin separated by 4 day intervals or left untreated (control). The results of this study demonstrated that the method of applying a suspension of oryzalin in warm, semi-solid agar to the shoots of Rhododendron seedlings was an effective method for inducing polyploidy. Although single applications resulted in some polyploid plants, multiple applications increased efficacy for some of the taxa studied. Treatments resulted in a range of ploidy levels, from 2x to 8x, including mixaploids (cytochimeras). The effect of increased ploidy level on pollen fertility and the occurrence of unreduced gametes in triploid taxa were also studied. Pollen viability was compared between corresponding progenitor and polyploid taxa by staining pollen with 1% acetocarmine (w⁄v) for 15 minutes. The existence of unreduced gametes in triploid taxa was determined by the presence of dyad and/or monad pollen grains. The results demonstrated that the fertility of polyploid Rhododendron can be highly variable and that the induction of polyploidy may either enhance or compromise fertility. Moreover, some triploids produced viable, unreduced pollen (as high as 5%), allowing for the possible utilization of these plants in breeding programs. Documentation of polyploid taxa, improved methods for inducing polyploidy, and information on fertility and reproductive biology of polyploid Rhododendron will be valuable to plant breeders. Methods for improving propagation of stem cutting of North American deciduous azaleas were also examined. Influence of dormant hedging, a range of rooting hormone concentrations (0, 2500, 5000, 7500, 10,000 ppm K-IBA), and tissue growth stage on rooting of Rhododendron austrinum (Small) Rehder and R. flammeum (Michx.) Sarg. were evaluated. Softwood stem cuttings of both species rooted from 70-90% while semi-hardwood cuttings rooted from 20-70% Cuttings from hedged stock plants rooted at higher percentages and possessed higher root system indexes compared to the unhedged counterparts in R. flammeum, but the effect of hedging was less evident in R. austrinum. Increasing IBA concentration increased rooting percentage of softwood cuttings of R. flammeum, but had no beneficial effect on rooting of semi-hardwood cuttings of R. flammeum or on R. austrinum, regardless of growth stage.



cutting propagation, polyploid induction, unreduced gametes, pollen staining, flow cytometry, rhododendron, allopolyploidy, autopolyploidy





Horticultural Science