Genetic diversity studies using 39 simple-sequence repeat (SSR) markers were carried out with 114 taxa of Hydrangea macrophylla (Thunb.) Ser., including 87 H. macrophylla ssp. macrophylla cultivars and 20 members of H. macrophylla ssp. serrata (Thunb.) Makino. The SSR loci were highly variable among the taxa, producing a mean of 8.26 alleles per locus. Overall allelic richness was relatively high at 5.12 alleles per locus. H. macrophylla ssp. serrata contained nearly twice the allelic diversity of H. macrophylla ssp. macrophylla. The majority of genetic diversity was found to reside within the subspecies, with only 12% of the total genetic diversity observed occurring between subspecies. Although the elevation of H. macrophylla ssp. serrata to species level has recently been recommended by several hydrangea authorities, these data support the subspecies designation. Four cultivars (Preziosa, Pink Beauty, Tokyo Delight, and Blue Deckle) appeared to be hybrids between the two subspecies. Genetic similarities were found among five remontant cultivars (Bailmer, Oak Hill, David Ramsey, Decatur Blue, and Penny Mac) and several nonremontant cultivars, including General Vicomtesse de Vibraye, Nikko Blue, All Summer Beauty, and La France. No close genetic relationship was found between the remontant cultivar Early Sensation and other remontant cultivars. Genetic similarities were found among variegated and double-flower cultivars. Within H. macrophylla ssp. macrophylla, cultivars with mophead inflorescences clustered separately from most lacecap cultivars. This indicates the cultivars with lacecap inflorescences that were among some of the earliest introductions to Europe were not widely used in the breeding of mophead forms. Some presumed synonyms were found to be valid (‘Preziosa’ and ‘Pink Beauty’, ‘Rosalba’ and ‘Benigaku’, ‘Geoffrey Chadbund’ and ‘Mowe’), whereas others were not (‘Harlequin’ and ‘Monrey’, ‘Nigra’ and ‘Mandschurica’). This study identified potentially unexploited sources of germplasm within H. macrophylla and relationships between existing cultivars of this popular shrub. This information should be of value when selecting parents for breeding programs.
Sandra M. Reed and Timothy A. Rinehart
Timothy Rinehart, Sandra Reed, and Brian Scheffler
Hydrangea popularity and use in the landscape has expanded rapidly in recent years with the addition of remontant varieties. Relatively little is known about the genetic background or combinability of these plants. We recently established microsatellite markers for hydrangea and evaluated their utility for estimating species diversity and identifying cultivars. We also verified an interspecific cross using these markers. Future research includes marker assisted breeding, particularly with respect to remontant flowering traits.
Sandra M. Reed and Timothy A. Rinehart
Genetic diversity studies using 26 simple-sequence repeat (SSR) markers were conducted with 36 cultivars, breeding lines, and wild-collected accessions of Hydrangea paniculata Sieb. The SSR markers were highly variable among the genotypes, producing a mean of 5.8 alleles per marker. Three cultivars (Boskoop, Compact Grandiflora, and Webb) were either identical to or sports of the popular cultivar Grandiflora. The name ‘Pee Wee’ appears to have been applied to two phenotypically different compact forms of H. paniculata, one of which seems to be a sport of ‘Tardiva’, whereas the other is likely derived from ‘Grandiflora’. No close genetic similarity was observed among several cultivars from a long-term Belgium breeding program, although many had one parent in common. Early-flowering genotypes clustered separately from genotypes that flower in midsummer, but close genetic relationships were not observed among early-flowering cultivars. Two genotypes from Taiwan were genetically similar but were distinctly different from the Japanese genotypes. These, along with the early-flowering genotypes and a new collection from Japan, may represent unexploited sources of germplasm for improvement of H. paniculata.
Stephen Patrick Greer and Timothy A. Rinehart
To determine the optimal conditions for mutagenesis of Hydrangea macrophylla (Thunb.) Ser. and Hydrangea paniculata Siebold, cold-treated and untreated seed from representative cultivars were exposed to 0.5%, 2.5%, and 5.0% ethyl methane sulfonate (EMS). Most untreated H. macrophylla seed exposed to 2.5% and 5.0% EMS had substantially lower germination percentages. H. paniculata seed germination percentages did not differ from controls until EMS concentrations reached 5.0%. Cold treatment of H. macrophylla and H. paniculata seed made germination more tolerant to all concentrations of EMS tested and increased germination of most H. macrophylla cultivars when compared with lower dosages. Germination of cold-treated seed from H. paniculata cultivars rose substantially above control levels even at the highest dosages. For the most part, we observed more nondormant seed, or seed available for germination, but less viability with increasing EMS dosage in untreated Hydrangea L. seed versus controls. In contrast, cold-treated seed displayed higher levels of mutagen tolerance and nondormancy versus controls when exposed to these same doses of EMS.
Keri D. Jones, Sandra M. Reed, and Timothy Rinehart
Dichroa febrifuga (Hydrangeaceae) is an evergreen shrub with metallic blue fruit that persists throughout winter. Phylogenetic studies have indicated that D. febrifuga is closely related to Hydrangea macrophylla. The objective of this study was to produce hybrids between these two ornamental species. Reciprocal crosses were made between D. febrifuga and both diploid and triploid cultivars of H. macrophylla. Two-thirds of the seed capsules produced from these hybridizations were allowed to mature on the plant, after which time seed were collected and sown in a greenhouse. Embryo rescue was used with the remaining seed capsules. Ovules were excised from ovaries about 9 weeks after pollination and plated on Gamborg's B-5 media with 2% sucrose. Both germination techniques resulted in vigorous seedlings. Hybridity was verified in plants obtained from ovule culture using simple sequence repeat (SSR) markers. Reciprocal hybrids were obtained using both diploid and triploid forms of H. macrophylla. This study confirms the close relationship between D. febrifuga and H. macrophylla and provides valuable material for future breeding efforts.
Timothy A. Rinehart, Brian E. Scheffler, and Sandra M. Reed
Using 14 codominant microsatellite markers that amplify loci across 14 different Hydrangea L. species, we analyzed gene diversity and genetic similarity within Hydrangea. Samples also included Dichroa Lour., Platycrater Sieb. and Zucc., and Schizophragma Sieb. and Zucc. genera to establish their relatedness to Hydrangea species since previous work suggests they may be closely related. Our results support the close affiliation between Macrophyllae E.M. McClint. and Petalanthe (Maxim.) Rehder subsections and their separation from the other Hydrangea species. Most of the Hydrangea species analyzed cluster within their designated sections and subsections; however, genetic distance between species within each subsection varied considerably. Our data suggest that morphological analyses which labeled H. serrata (Thunb.) Ser. as a subspecies of H. macrophylla (Thunb. Ex J.A. Murr.) Ser. are probably more accurate than recent genome size data suggesting H. macrophylla ssp. macrophylla (Thunb.) Ser. and H. macrophylla ssp. serrata (Thunb.) Makino are separate species. Gene diversity estimates indicate that 64.7% of the total diversity is due to differences between species and 49.7% of the overall variation is due to differences between subsections. Low diversity suggests a lack of gene flow between species and subsections and underscores the difficulty in making wide hybrids. Since only 35.3% of the genetic variation is common to all species, unique alleles were used to develop a molecular key for unambiguous species identification and interspecific hybrid verification. Genetic similarity estimates for all 85 samples suggests a roadmap for introgressing horticulturally important traits from different Hydrangea species.
Sandra M. Reed, Keri D. Jones, and Timothy A. Rinehart
The potential of producing an intergeneric hybrid between Dichroa febrifuga Lour. and Hydrangea macrophylla (Thunb.) Ser. was investigated. Reciprocal hybridizations were made between a D. febrifuga selection (GUIZ 48) and diploid (‘Veitchii’) and triploid (‘Kardinal’ and ‘Taube’) cultivars of H. macrophylla. Embryo rescue was employed for about one-third of the crosses that produced fruit, and the rest were allowed to mature on the plant and seed-collected and germinated. Reciprocal hybrids, which were verified with simple sequence repeat markers, were produced from both embryo rescue and seed germination and with both diploid and triploid H. macrophylla cultivars. Hybrids were intermediate in appearance between parents, but variability in leaf, inflorescence, and flower size and flower color existed among the hybrids. A somatic chromosome number of 2n = 6x = 108 was tentatively proposed for D. febrifuga GUIZ 48. Chromosome counts and flow-cytometric measurements of nuclear DNA content indicated that some of the hybrids may be aneuploids, but neither analysis was definitive. Although hybrids with H. macrophylla as the pistillate parent did not form pollen-producing anthers, D. febrifuga × H. macrophylla hybrids had normal-appearing anthers that produced abundant pollen. F2 and BC1 progeny were obtained using D. febrifuga × ‘Veitchii’ hybrids. This work documents the first step in an effort to combine desirable horticultural features from D. febrifuga and H. macrophylla.
Timothy A. Rinehart, Ned C. Edwards, and James M. Spiers
Keri D. Jones, Sandra M. Reed, and Timothy A. Rinehart
Ploidy level was estimated in Hydrangea macrophylla (Thunb) Ser. using flow cytometry. For H. macrophylla ssp. macrophylla, 42 diploid and 19 triploid cultivars were identified. All 14 H. macrophylla ssp. serrata (Thunb.) Makino cultivars tested were diploids. Somatic chromosome counts confirmed the ploidy of three diploid (2n = 2x = 36) and three triploid (2n = 3x = 54) cultivars. Stomatal guard cell length and pollen diameter of H. macrophylla ssp. macrophylla diploid cultivars were smaller than those of triploid cultivars. However, because the range of measurements for the diploids overlapped that of the triploids, neither guard cell nor pollen measurements are recommended for determining ploidy of H. macrophylla cultivars. Fertility was estimated using pollen staining and controlled pollinations. Stainable pollen for triploid cultivars averaged 63% and ranged from 25% in ‘Masja’ to 85% in ‘Marechal Foch’. Viable seed was obtained when four triploid cultivars were used as pistillate or staminate parents in controlled pollinations to diploid H. macrophylla ssp. macrophylla cultivars. A bimodal distribution of pollen sizes, which is suggestive of unreduced gamete production, was observed in one cultivar; however, more detailed genetic and cytologic studies are needed to elucidate the mechanism behind triploid formation in H. macrophylla taxa.