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Hui Cao, Hui Cao, Dennis B. McConnell and Jianjun Chen*

The irritant effect of Dieffenbachia sap is attributed to protelytic enxymes but calcium oxalate crystals are considered to puncture cells and allow enzyme entrance. To date, no detailed study of the location, type, or frequency of calcium oxalate crystals in Dieffenbachia species or cultivars has been undertaken. To do so, three uniform tissue culture plantlets of Dieffenbachia `Carina',`Rebecca' or `Star Bright' were transpanted into 15 cm pots, grown in a shaded greenhouse under 385 μmol·m-2·s-1 and fertigated with 20 N-8.7 P-16.6 K water-soluble fertilizer at N concentrations of 200 mg·L-1 twice weekly. Ten weeks later, samples of stem, root, and leaves were taken from 4 pots of each cultivar to determine the distribution and type of calcuium oxalate crystals in each plant organ via polarized light microscopy. Two types of calcium oxlate crystals, raphides and druses, were found in the stem, leaves and roots. Druse density increased as leaves andd stems matured while the number of raphide idioblasts remained relatively constant. Crystal density was highest at lateral initation sites of buds and roots. Significant differences were found in crystal density among cultivars even though `Carina' and `Star Bright' are sports selected from `Camille'. This suggests that reduction of calcium oxalate density of Dieffenbachia cultivars is possible through breeding.

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Richard J. Henny, James R. Holm, Jianjun Chen and Michelle Scheiber

Colchicine application successfully induced tetraploids from in vitro-cultured diploid Dieffenbachia × ‘Star Bright M-1’. Shoot clumps, each with six to eight small, undifferentiated shoot primordia, were cultured in liquid Murashige and Skoog (MS) medium and treated with colchicine at rates of 0, 250, 500, or 1000 mg·L−1 for 24 h. In vitro survival of shoot clumps significantly decreased as colchicine concentrations increased. Shoot clumps that survived were transferred to colchicine-free MS medium containing 2.0 mg·L−1 N6-isopentenyl) adenine and 0.10 mg·L−1 indole-3-acetic acid. Shoots were harvested during four subsequent subcultures and planted in a soilless substrate in a shaded greenhouse. The number of plants that survived 6 months after ex vitro planting was 690, 204, 59, and 69 for colchicine treatments at 0, 250, 500, and 1000 mg·L−1, respectively. The 332 plants from colchicine treatments along with 90 control plants (selected from 690 in the control treatment) were evaluated morphologically in a shaded greenhouse. Overall plant growth, including crown height, plant canopy, and leaf size, of colchicine-treated plants was significantly less than controls. Based on the growth data, 10, 32, 15, and 16 plants from the 0, 250, 500, and 1000 mg·L−1 colchicine rates, respectively, were selected and analyzed by flow cytometry. Flow cytometry confirmed the presence of 13 tetraploids and 29 mixoploids among the 63 colchicine-treated selections; all 10 plants from the control were diploid. A colchicine rate of 500 mg·L−1 produced a higher percentage of tetraploids (10.2%) than did the 250 (2.9%) or 1000 mg·L−1 (1.4%) rates. Subsequent comparisons showed tetraploids had significantly smaller and thicker leaves, greater specific leaf weights, and longer stomata than diploids. Tetraploids also showed increased net photosynthetic rate, decreased g S, decreased intercellular CO2 concentration, decreased transpiration rate, and increased water use efficiency. Tetraploids appeared robust and their smaller size could make them potentially more durable plants used as living specimens for interior decoration.

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Xiaoming Wang, Jianjun Chen, Yongxin Li, Qiying Nie and Junbin Li

‘Jincuilei’ is a mutant selected from Lonicera macranthoides Hand.-Mazz. It produces abundant flowers that never open with a chlorogenic acid (CGA) content up to 6.0%. Propagation through rooting or grafting has only a 30% survival rate. This study was undertaken to establish an efficient protocol for rapidly regenerating this mutant. Leaf explants were inoculated on Gamborg's B5 medium supplemented with different concentrations of 6-benzyladenine (BA) and 2,4-dichlorophenozyacetic acid (2,4-D). The optimal combination for callus induction was 4.4 μm BA with 2.26 μm 2,4-D, which resulted in 86.7% of leaf explants producing calluses in 4 weeks. Calluses produced from this optimal medium were cultured on B5 medium containing different concentrations of kinetin (KT) and α-naphthalene acetic acid (NAA). The best formulation for shoot induction was B5 medium containing 0.9 μm KT and 5.4 μm NAA in which 73.4% of cultured calluses produced shoots in 8 weeks, and shoot numbers ranged from three to six per callus piece (1 cm3). Adventitious shoots were cut and rooted in half-strength Murashige and Skoog medium supplemented with 14.8 μm 3-indolebutyric acid. Roots initiated 10 d after culture, and rooting percentages ranged from 98% to 100%. Plantlets grown in a container substrate in a shaded greenhouse had over a 95% survival rate. During the last 6 years, over four million plantlets were regenerated using this established procedure, and there was no somaclonal variation. Fresh and dry weights of 1000 flowers, CGA contents, and dry flower yields of the regenerated plants were not significantly different from those of the stock ‘Jincuilei’ propagated by cutting, indicating that plants regenerated from this established procedure were stable. This established in vitro culture method has led to rapid commercial production of this medicinal plant on more than 1500 ha of production field.

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Richard J. Henny, Jianjun Chen and Terri A. Mellich

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Min Deng, Jianjun Chen, Richard J. Henny and Qiansheng Li

Codiaeum variegatum (L.) Blume is one of the most popular ornamental foliage plants. It encompasses more than 300 recognized cultivars valued by their wide range of leaf shapes and vivid foliage colors. Thus far, only limited information is available regarding the genetic basis of their leaf morphological variation. This study investigated the chromosome numbers and karyotypes of seven phenotypically diverse cultivars. Root-tip cells were fixed, mounted, and observed under light microscopy. Results showed that chromosome numbers in the mitotic metaphase of the seven cultivars were high and variable and ranged from 2n = 66, 70, 72, 76, 80, 82, 84, to 2n = 96, indicating that the cultivars are polyploid and some could be aneuploid. Genetic mosaics occurred in one of the seven cultivars. Additionally, each cultivar had its own karyotype. There were no relationships between chromosome numbers or karyotypes and leaf morphology. Results from this study suggest that the morphological diversity among cultivars of this species could be in part attributed to high variation in chromosome numbers and karyotypes.

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Min Deng, Jianjun Chen, Richard J. Henny and Qiansheng Li

Codiaeum variegatum (L.) Blume, commonly known as crotons, are among the most popular ornamental foliage plants cultivated for either landscaping or interiorscaping. Currently, more than 300 cultivars are available; each has a distinct phenotype, particularly in leaf morphology. Thus far, there is no information regarding their genetic relationships. In this study, genetic relatedness of 44 cultivars of C. variegatum was investigated using amplified fragment length polymorphism (AFLP) markers. Fourteen primer combinations generated a total of 549 AFLP fragments, which were used to estimate genetic distances and construct dendrograms based on the neighbor-joining method. The 44 cultivars were divided into seven clusters, which concurred with the known history of croton geographical isolation, adaptation, introduction, and breeding activities but differed from the classification made by the Croton Society based on leaf morphology. The established genetic relationships could be important for future germplasm identification and conservation and new cultivar development. Additionally, genetic distance among the 44 cultivars was 0.322 or less, indicating that they have a narrow genetic base. The narrow genetic base may indicate that the cultivars were derived from a common progenitor. On the other hand, 81% of the 549 fragments were polymorphic and the average polymorphic information content was 0.22, which suggests that the cultivars are genetically highly polymorphic. The high polymorphisms may be attributed to significant gene loss or gain facilitated by mutation and/or chromosome variation, thus contributing to a wide range of leaf morphological differences among cultivars.

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Xiaoying Li, Hongxia Xu, Jianjun Feng and Junwei Chen

Deep transcriptome sequencing allows for the acquisition of large-scale microsatellite information, and it is especially useful for genetic diversity analysis and mapping in plants without reference genome sequences. In this study, a total of 14,004 simple sequence repeats (SSRs) were mined from 10,511 unigenes screening of 63,608 nonredundant transcriptome unigenes in loquat (Eriobotrya japonica) with a frequency of 22 SSR loci distributed over 100 unigenes. Dinucleotide and trinucleotide repeat SSRs were dominant, accounting for 20.62%, and 42.1% of the total, respectively. Seventy primer pairs were designed from partial SSRs and used for polymerase chain reaction (PCR) amplification. Of these primer pairs, 54 exhibited amplification and 33 were polymorphic. The number of alleles at these loci ranged from two to 17, and the polymorphism information content values ranged from 0.24 to 0.89. We tested the transferability of 33 SSR polymorphic primer pairs in apple and pear, and the transferability rates in these two species were 90.9% and 87.9%, respectively. A high level of marker polymorphism was observed in apple [Malus ×domestica (66.7%)], whereas a low level was observed in pear [Pyrus sp. (51.5%)]. In addition, the PCR products from seven SSR primer pairs were selected for sequence analysis, and 89.2% of the fragments were found to contain SSRs. SSR motifs were conserved among loquat, apple, and pear. According to our sequencing results for real SSR loci, ≈12,490 SSR loci were present in these loquat unigenes. The cluster dendrogram showed a distinct separation into different groups for these three species, indicating that these SSR markers were useful in the evaluation of genetic relationships and diversity between and within the species of Maloideae in the Rosaceae. The results of our identified SSRs should be useful for genetic linkage map construction, quantitative trait locus mapping, and molecular marker-assisted breeding of loquat and related species.

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Kaitlyn McBride, Richard J. Henny, Jianjun Chen and Terri A. Mellich

Adenium obesum (Forssk.), Roem. & Schult., commonly known as desert rose, is a high-value, container-grown ornamental plant produced worldwide for its bright floral display and striking sculptural caudex. Little scientific-based information exists regarding the effect of light intensity and nutritional levels on Adenium growth and flowering. In this study, A. obesum ‘Red’ and ‘Ice Pink’ were grown under full sun [with a measured maximum photosynthetically active radiation (PAR) of 1850 μmol·m−2·s−1], 30% shade (1255 μmol·m−2·s−1), or 50% shade (943 μmol·m−2·s−1) in 1.25-L pots top-dressed with controlled-release fertilizer Nutricote® Plus (18N–2.6P–6.6K) at rates to provide 0.4, 0.9, or 1.4 g of nitrogen (N) per pot. Canopy height and width, flower number, and visual quality ratings (based on plant size and form, foliage color, and flowering) were highest after 16 weeks of growth for both cultivars when fertilized with 1.4 g of N per pot. A 30% shade level resulted in plants with the highest flower numbers and quality ratings. Plants grown at 50% shade had the greatest canopy heights and widths, but flower numbers and quality ratings were low. In full sun, plants were smaller overall. In a second experiment, A. obesum ‘Red’ produced the highest shoot dry weight when grown 20 weeks at 30% or 50% shade with 1.4 g of N per pot. Root formation is an important measure of aesthetic value for this crop. As plants mature, roots enlarge dramatically and are often washed to expose sculptural forms. The highest root dry weights were measured at 1.4 g of N under both full sun and 30% shade.

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Qiansheng Li, Jianjun Chen, Dennis B. McConnell and Richard J. Henny

A simple and effective method for quantification of leaf variegation was developed. Using a digital camera or a scanner, the image of a variegated leaf was imported into a computer and saved to a file. Total pixels of the entire leaf area and total pixels of each color within the leaf were determined using an Adobe Photoshop graphics editor. Thus, the percentage of each color's total pixel count in relation to the total pixel count of the entire leaf was obtained. Total leaf area was measured through a leaf area meter; the exact area of this color was calculated in reference to the pixel percentage obtained from Photoshop. Using this method, variegated leaves of ‘Mary Ann’ aglaonema (Aglaonema x), ‘Ornate’ calathea (Calathea ornate), ‘Yellow Petra’ codiaeum (Codiaeum variegatum), ‘Florida Beauty’ dracaena (Dracaena surculosa), ‘Camille’ dieffenbachia (Dieffenbachia maculata), and ‘Triostar’ stromanthe (Stromanthe sanguinea) were quantified. After a brief training period, this method was used by five randomly selected individuals to quantify the variegation of the same set of leaves. The results were highly reproducible no matter who performed the quantification. This method, which the authors have chosen to call the quantification of leaf variegation (QLV) method, can be used for monitoring changes in colors and variegation patterns incited by abiotic and biotic stresses as well as quantifying differences in variegation patterns of plants developed in breeding programs.

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Jianjun Chen, Russell D. Caldwell and Cynthia A. Robinson

Gynura aurantiaca is a colorful foliage plant with creeping stems and velvety purple hairs that cover the green leaves. It grows rapidly, but is cultivated primarily for those attractive purple leaves. Annually during the spring, this plant produces prominent flowers both in appearance and smell, gaudy and malodorous. Flowering coupled with acquiring an over-grown leggy appearance have been key limitations in its production and use in interiorscaping. This study was undertaken to determine if an available commercial plant growth regulator could inhibit flowering. A-Rest (ancymidol), B-Nine (daminozide), Bonzi (paclobutrazol), cycocel (chlormequat chloride) and florel (ethephon) each diluted to three different concentrations were sprayed in two applications in early spring at 2-week intervals. Flowering and bud numbers and plant growth (number of lateral shoots, vine lengths and internode lengths) were recorded. Results indicated that applications of A-Rest, B-Nine, Bonzi and Cycocel, regardless of treatment concentrations, were ineffective in suppressing the flowering of this plant; whereas, florel completely suppressed flowering at the three concentrations used. The florel-treated plants also grew more lateral shoots, which produced a compact and dense bush-look, indicating that appropriate concentrations of florel application not only will stop flowering of purple passion but can also improve and prolong its aesthetic value as a potted or hanging-basket interior plant.