Quantification of phenolic acids and antioxidant activity in sweetpotato genotypes

Phenolic acid composition and antioxidant activity in roots of 14 commercially important sweetpotato genotypes were evaluated. Significant differences in total phenolics, individual phenolic acids, and antioxidant activity were found among the different sweetpotato genotypes. Total phenolic content, expressed in terms of chlorogenic acid equivalent, in different genotypes ranged from 1.4 to 4.7 mg g⁻¹ dry weight (DW). Antioxidant activity was evaluated as Trolox equivalent, ranging from 1.3 to 4.6 mg g⁻¹ DW. The highest total phenolic content and antioxidant activity were observed in a purple-fleshed genotype. Chlorogenic acid and 3,5-dicaffeoylquinic acid were the predominant phenolic acids, while caffeic acid was the least abundant in most genotypes. The highest content of chlorogenic acid (422.4 μg g⁻¹ DW) was present in a white-fleshed cultivar ‘Quarter Million’ imported from Jamaica. However, a purple-fleshed genotype had the highest amounts of 3,5-dicaffeoylquinic (485.6 μg g⁻¹ DW), 3,4-dicaffeoylquinic (125.6 μg g⁻¹ DW), 4,5-dicaffeoylquinic (284.4 μg g⁻¹ DW), and caffeic (20.5 μg g⁻¹ DW) acids.     

Plant regeneration of pansy (Viola wittrockiana) ‘Caidie’ via petiole-derived callus

An in vitro plant regeneration protocol for pansy (Viola wittrockiana) cultivar ‘Caidie’ from petioles was established as following: callus induction on a half-strength MS medium supplemented with 0.45 μmol l⁻¹ 2,4-d plus 8.9 μmol l⁻¹ BA, callus subculture on medium F (1/2MS with 4.5 μmol l⁻¹ 2,4-d, 2.7 μmol l⁻¹ NAA and 0.44 μmol l⁻¹ BA) and then on medium T (1/2MS with 4.5 μmol l⁻¹ 2,4-d, 2.7 μmol l⁻¹ NAA and 2.2 μmol l⁻¹ BA), shoot regeneration on medium D3 (MS media supplemented with 2.9 μmol l⁻¹GA₃, 23.6 μmol l⁻¹ AgNO₃, 0.02% active charcoal and 4.5 μmol l⁻¹ TDZ), shoot multiplication on medium M (half-strength MS medium containing NAA 1.1 μmol l⁻¹, TDZ 9.1 μmol l⁻¹ and GA₃ 8.7 μmol l⁻¹), and then shoot elongation and rooting on medium R (MS medium supplemented with 1.1 μmol l⁻¹ NAA and 1.1 μmol l⁻¹ BA). Subculture on appropriate medium was found to be important for successful shoot regeneration.     

Temperature and photoperiod control of morphology and flowering time in two greenhouse grown Hydrangea macrophylla cultivars

Knowledge of the factors involved, and tools to control morphology and flowering are important in intensive and cost-efficient greenhouse production. Hydrangea macrophylla is an important flowering pot plant in Norway and is produced year-around in greenhouses. Due to problems in scheduling, a study was conducted to compare floral transition and morphology of two commercially important cultivars of Hydrangea (‘Early Blue’ and ‘Schneeball’) under different flower initiating treatments in growth chambers. Plants were grown with high pressure sodium lamps (HPS) at moderate temperature (17 °C) (MT) and high (24 °C) temperature. At high temperature, the effect of (1) irradiance under long day conditions (16 h lighting with 70 or 200 μmol m⁻² s⁻¹), and (2) short day (8 h lighting) was investigated. The short day treatment had similar light integral as the low irradiance long day treatment (SD: 8 h × 140 μmol m⁻² s⁻¹ and LD: 16 h × 70 μmol m⁻² s⁻¹ = 4.0 mol m⁻² d⁻¹). The intention was to test the effect of irradiance and SD on flower transition and morphology under high temperatures. The results clearly showed that MT is the strongest signal for floral transition. MT resulted in a rapid floral transition of the terminal buds and lateral flower buds. A short forcing period was required and the plants became short and compact without any use of chemical growth retardants. At high temperatures only SD had a promotive effect on flower transition and the response was found to be stronger in ‘Schneeball’ than ‘Early Blue’. In general, all the treatments under high temperatures required a long forcing time and the plants tended to be very tall with a low number of lateral flower buds.     

Growth and photosynthetic characteristics of blueberry (Vaccinium corymbosum cv. Bluecrop) under various shade levels

Blueberry can readily be shaded as a bush type plant, maybe affecting its growth and photosynthesis. Growth and photosynthetic characteristics of ‘Bluecrop’ blueberry grown under various shade levels were investigated to understand acclimation under shade conditions and to determine the optimal light conditions for agricultural purpose. Shade decreased the number of shoots per shrub, but increased shoot length. However, shade did not affect the number of leaves on the main axis. With increasing shade level, leaf length, width and area increased, but leaf thickness decreased. However, there was no obvious tendency in leaf length/width ratio with increasing shade level. Shade leaves had less dense stomata than sun leaves, but stoma was bigger in shade leaves than in sun leaves. With increasing shade level, non-photochemical quenching in blueberry leaves increased and the values were higher at low photosynthetic photon flux densities (PPFDs) in shade leaves than in sun leaves, resulting in the decreases in quantum yield, electron transport rate and net CO₂ assimilation rate (A_n). The maximum A_n at 31, 60, 73 and 83% shade levels was 11.8, 11.0, 8.4 and 7.5 μmol m⁻² s⁻¹, respectively. Following the slight decrease up to 100 μmol m⁻² s⁻¹ PPFD, stomatal conductance (g_s) linearly increased up to 600 μmol m⁻² s⁻¹ PPFD and became saturated at all shade levels. The leaves of the shrubs grown under the 83% shade level had a significantly lower g_s as compared to the leaves of the shrubs grown under the 31, 60 and 73% shade levels. Transpiration rate (E) linearly increased up to 600 μmol m⁻² s⁻¹ PPFD and was saturated at the 73 and 83% shade levels. However, E increased linearly at both 31 and 60% shade levels with increasing PPFD. The reproductive growth characteristics such as number of flowers, fruit set rate per flower bud and fruit yield also significantly decreased with increasing shade level. For agricultural purpose, therefore, shade level above approximately 60% of full sunlight must be avoided for optimal photosynthesis and growth of the ‘Bluecrop’ blueberry.     

Night interruption promotes vegetative growth and flowering of Cymbidium

The effects of night interruption (NI) were examined on the vegetative growth and flowering of Cymbidium ‘Red Fire’ and ‘Yokihi’. Plants were grown under 9/15 h ambient light/dark (control), 9 h ambient light plus night interruption (22:00–02:00 h) with low light intensity at 3–7 μmol m⁻² s⁻¹ (LNI) and 9 h ambient light plus NI with high light intensity at 120 μmol m⁻² s⁻¹ (HNI) conditions. The number of leaves, leaf length, number of pseudobulbs and pseudobulb diameter increased in both LNI and HNI compared to controls for both cultivars. While none of the control plants flowered within 2 years, 100% of the ‘Yokihi’ and 80% of the ‘Red Fire’ plants grown under HNI condition flowered. In the LNI group, 60% of the plants flowered in both cultivars. Plants in the HNI group showed a decreased time to visible inflorescence and flowering than those in the LNI group. The number of inflorescences and florets were greater in the plants grown under HNI than those in the LNI group. The tallest plants at flowering were in the HNI group in both cultivars. NI with low light intensity can be used effectively to promote flower induction with increased growth rate during the juvenile stage in Cymbidium. To obtain high quality plants, however, NI with high light intensity strategies should be considered.     

Modeling plant morphology and development of petunia in response to temperature and photosynthetic daily light integral

The effects of mean daily temperature (MDT) and mean photosynthetic daily light integral (MDLI) on flowering during the finish stage of two petunia (Petunia × hybrida) cultivars were quantified. Petunia ‘Easy Wave Coral Reef’ and ‘Wave Purple’ were grown in glass-glazed greenhouses at 14–23 °C or 14–26 °C and under 4–19 mol m⁻² d⁻¹ with a 16-h photoperiod. The flower developmental rate was predicted using a model that included a linear MDT function with a base temperature multiplied by an exponential MDLI saturation function. The flower developmental rate increased and time to flower decreased as MDT increased within the temperature range studied. For example, under a MDLI of 12 mol m⁻² d⁻¹, as MDT increased from 14 to 23 °C, time to flower of ‘Easy Wave Coral Reef’ and ‘Wave Purple’ decreased from 51 to 22 d and 62 to 30 d, respectively. Flower developmental rate increased as MDLI increased until saturation at 14.1–14.4 mol m⁻² d⁻¹. Nonlinear models were generated for effects of MDT and MDLI on flower bud number and plant height at flowering. The number of flower buds at flowering increased as MDT decreased and MDLI increased. For example, at an MDT of 14 °C with 18 mol m⁻² d⁻¹, plants had 2.5–2.9 times more flower buds than those grown at 23 °C and 4 mol m⁻² d⁻¹. Models were validated with an independent data set, and the predicted time to flower, flower bud number, and plant height were within ±7 d, ±20 flowers, and ±4 cm, respectively, for 96–100%, 62–87%, and 93–100% of the observations, respectively. The models could be used during greenhouse crop production to improve scheduling and predict plant quality of these petunia cultivars.     

Antioxidant properties of blackberry and blueberry fruits grown in the Black Sea Region of Turkey

Seven wild and ten cultivated blackberries (Arapaho, Bartin, Black Satin, Bursa 1, Bursa 2, Cherokee, Chester, Jumbo, Navaho, and Ness), and six lowbush (Vaccinium arctostaphylos) and four highbush (Vaccinium corymbosum) blueberries fruits (Ivanhoe, Jersey, Northland, and Rekord) were analyzed for total anthocyanins, total phenolics, and antioxidant activity as ferric reducing antioxidant power (FRAP) in this study. The respective ranges of total anthocyanin and total phenolic contents of the tested samples were: blackberries, 0.95–1.97 and 1.73–3.79 mg g⁻¹ and blueberries, 0.18–2.94 and 0.77–5.42 mg g⁻¹. FRAP values varied from 35.05 to 70.41 μmol g⁻¹ for blackberries, 7.41 to 57.92 μmol g⁻¹ for blueberries. Wild blackberries had the highest FRAP values while wild blueberries had the highest total phenolic and total anthocyanin contents. A linear relationship was observed between FRAP values and total phenolics for blueberries (r = 0.981). The anthocyanin pigments in samples were isolated and characterized by high-performance liquid chromatography (HPLC) with UV–visible detection. Cyn-3-glu was the predominant anthocyanin in all blackberry fruits.     

Priming abiotic factors for optimal hybrid Cymbidium (Orchidaceae) PLB and callus induction,plantlet formation,and their subsequent cytogenetic stability analysis

Abiotic factors affect the induction of PLBs and callus in hybrid Cymbidium Twilight Moon ‘Day Light’. The initiation and proliferation of new PLBs and callus could be achieved on NAA and kinetin, supplemented at 0.1 mg l⁻¹ each, respectively, both within 45–60 days. Bacto agar was found to be the most suitable solidifying agent for PLB induction, although a higher shoot fresh weight was obtained on Gelrite; a pH 5.3 was optimal while pH 4.5 caused 100% explant necrosis; coconut water, when supplied at 10–20% (v/v) resulted in a significant increase in the number of PLBs formed per PLB segment (23.1 versus 14.6 in controls) while a massive (almost four-fold) increase in fresh top weight occurred when PLB explants were placed in liquid culture, as a result of hyperhydricity; Fe-EDTA (1 mg l⁻¹) and activated charcoal (1 g l⁻¹) stimulated total fresh weight and PLB formation in the presence of PGRs; PLB formation decreased but total fresh shoot weight increased with the addition of niacin or myo-inositol, both vitamins. Dark-grown PLB-induced plants were etiolated and had longer internodes and higher fresh weight than light-grown control plants at 45 μmol m⁻² s⁻¹; at 15 μmol m⁻² s⁻¹ shoots were slightly etiolated, fragile, and PLB formation was scarce. RAPD and mtDNA analysis of all resultant PLBs, callus or plants showed them to be genetically identical, with comparable chlorophyll contents. Despite the detection of cytological variation between different plant parts, little variation resulted from abiotic factor treatment.     

Chemical changes during myrtle (Myrtus communis L.) fruit development and ripening

The chemical composition of fruit belonging to ‘Barbara’ and ‘Daniela’ myrtle cultivars was monitored during development from fruit-set to an over-ripe stage (July–January), with the aim to identify a reliable maturity index. Acidity, pH, reducing and total sugars, phenols, tannins, anthocyans, carbon dioxide and ethylene production rates were monitored over two different year seasons. Titratable acidity decreased during maturation, with significant differences due to cultivar and year of observation. Reducing sugars increased in both cultivars approximately sevenfold from fruit set to complete maturation. Total sugar content increased similarly ranging from 1.43% and 1.41% at fruit set to 8.28% and 7.56% at maturation for ‘Barbara’ and ‘Daniela’, respectively. Total phenols and tannins occurred at high levels after fruit set and declined during development. Anthocyans levels increased, in both cultivars, according to a sigmoid curve. The pattern of respiration rate showed a gradual decline in both cultivars ranging from 365.81 and 396.42 mg kg⁻¹ h⁻¹ to 79.98 and 52.27 mg kg⁻¹ h⁻¹, respectively for ‘Barbara’ and ‘Daniela’ in 2006. A peak of variable size was observed in October–November period. Small increases in ethylene production have been detected during fruit development ranging from 130.57 and 269.14 μL kg⁻¹ h⁻¹ measured at the onset of development to 13.04 and 19.36 μL kg⁻¹ h⁻¹ measured at harvest for ‘Barbara’ and ‘Daniela’, respectively.     

Responses of eight chile peppers to saline water irrigation

Salt tolerance of five cultivars of Capsicum annuum L. Early Jalapeno, Golden Treasure, NuMex Sweet, NuMex Joe E. Parker, and Santa Fe Grande, two cultivars of C. chinense Jacq. Habanero and Pimienta De Chiera, and one accession of C. annuum, NMCA 10652, were evaluated in a field study. Seedlings were transplanted in late May to field raised beds containing loamy sand soils in a semi-arid environment. Plants were well irrigated throughout the experiment. Three saline solution treatments, prepared by adding NaCl, MgSO₄, and CaCl₂ to tap water at different amounts to create three salinity levels of 0.82 dS m⁻¹ (control, tap water), 2.5 dS m⁻¹, and 4.1 dS m⁻¹ electrical conductivity (EC), were initiated on 15th June and ended in late August. Among the eight varieties, NMCA 10652 had the highest survival percentage at 100% in the 4.1 dS m⁻¹ treatment, followed by ‘Early Jalapeno’, ‘NuMex Sweet’, ‘Pimienta De Chiera’, ‘Santa Fe Grande’, ‘Golden Treasure’, and ‘NuMex Joe E. Parker’. ‘Habanero’ had the lowest survival at 28%. Compared to control, final shoot dry weight of the plants irrigated with saline solution at 4.1 dS m⁻¹ was reduced by 92% in ‘Habanero’, followed by ‘Golden Treasure’ at 80%. For fruit fresh weight in 4.1 dS m⁻¹ vs. control, ‘Habanero’ had the highest reduction at 86%, followed by ‘Golden Treasure’ at 74%, while NMCA 10652 and ‘Santa Fe Grande’ had the least at 26% and 19%, respectively. NMCA 10652, the most tolerant to salinity, had the lowest leaf Na⁺ accumulation, while ‘Habanero’, the most sensitive to salinity, had the highest Na⁺ in the leaves. For leaf Cl⁻, ‘Early Jalapeno’ had the highest, while ‘Habanero’ had the lowest Cl⁻ accumulation in the leaves. Generally, sensitive varieties accumulated more Na⁺ and/or Cl⁻ in leaves, except for ‘Early Jalapeno’, which was relatively tolerant to salinity but had high Na⁺ and Cl⁻ accumulation in leaves.     

Analysis and modeling of gas exchange processes in Scaevola aemula

Scaevola aemula is a popular ornamental crop cultivated as a bedding plant or for hanging baskets. We characterized gas exchange properties of S. aemula ‘New Wonder’ in response to photosynthetically active radiation (PAR), carbon dioxide concentration, and leaf temperature. Net CO₂ assimilation rate (A) was responsive to CO₂, exhibiting a saturation when intercellular CO₂ concentration (C_i) was greater than 600 μmol mol⁻¹. Net CO₂ assimilation rate and dark respiration rate (R_d) were 23.1 and 2.3 μmol m⁻² s⁻¹, respectively, at 25 °C and PAR = 1500 μmol m⁻² s⁻¹. Net CO₂ assimilation rates were similar at leaf temperatures between 20 and 30 °C but significantly reduced at 15 °C. These gas exchange results were used to test the extendibility of a coupled gas exchange model previously developed for cut-roses. Utilizing the gas exchange data measured at 25 °C leaf temperature, several model parameters were independently determined for S. aemula. Model predictions were then compared with observations at different leaf temperatures. The model predicted the rates of net CO₂ assimilation and transpiration of S. aemula reasonably well. Without additional calibration, the model was capable of predicting the temperature dependence of net CO₂ assimilation and transpiration rates. Applying the model to predict the effects of supplemental lighting and CO₂ enrichment on canopy photosynthesis and transpiration rates, we show that this model could be a useful tool for examining environmental control options for S. aemula production in the greenhouse.     

Effects of low temperature storage on growth and transplant quality of non-grafted and grafted cantaloupe-type muskmelon seedlings

Grafting is a unique horticultural technology that allows the grower to select an alternate, compatible root system with natural disease resistance for their desired crop. Short-term storage of grafted seedlings under low temperature may extend the production window of grafted seedlings, reduce the labor input and increase production of grafted seedlings with a small propagation capacity. To evaluate the low temperature storage conditions, Cucumis melo ‘Olympic Gold’ seedlings were grafted onto Cucurbita maxima x Cucurbita moschata ‘Tetsukabuto’ rootstock and stored for a period of 2 or 4 weeks at 9, 12, or 15 °C under 12 μmol m⁻² s⁻¹ photosynthetic photon flux (PPF). The study demonstrated that grafted seedlings could be stored at 12 °C for 4 weeks without significant dry mass accumulation or effects on post-storage growth and development. Grafted seedlings stored at 15 °C for 4 weeks had a significant increase in dry mass and stem elongation; this was not observed for the non-grafted seedlings stored under the same conditions, suggesting that the rootstock enhanced the scion growth at lower temperatures than optimal for muskmelon. Storing muskmelon seedlings at 9 °C caused chilling damage but the damage was pronounced for non-grafted seedlings than grafted seedlings. ‘Tetsukabuto’ rootstock, an interspecific squash, presumably has a chilling tolerance and increased the storability of muskmelon seedlings. Further optimization is needed but there is potential for using this technique as a tool for mass production of grafted muskmelon seedlings.     

Involvement of proline in response of chickpea (Cicer arietinum L.) to chilling stress at reproductive stage

Chickpea is sensitive to chilling stress, especially at its reproductive stage and experiences abortion of flowers and poor pod set at temperatures below 10 °C. The metabolic controls governing chilling-sensitivity in chickpea, particularly involving proline are not known. Hence, in the present study we explored the role of proline in this regard. A set of chickpea plants (cv. GPF2) growing under warm conditions of the glass house (temperature – 28/14 °C as average maximum and minimum till early flowering stage) was exposed to low temperature conditions of the field (8.3–9.6/2.8–5.3 °C; average maximum and minimum temperature, respectively) during the onset of reproductive phase while another set of plants continued to grow under warm conditions. In case of chilling-stressed plants, one set of the plants was treated with 10 μM proline while the other set not treated with proline served as control under low temperature conditions. In untreated chilling-stressed plants, the endogenous proline increased to 230 μmol g⁻¹ dry weight (DW) on 4th day of stress and decreased thereafter to reach 28 μmol on 7th day. In plants treated with 10 μM proline, its endogenous content reached 310 μmol g⁻¹ DW on 4th day and stayed significantly higher than untreated chilling-stressed plants. The proline-treated plants showed significant improvement in retention of flowers and pods leading to better seed yield compared to the untreated ones. The proline-applied plants also had greater pollen viability, pollen germination, pollen tube growth and ovule viability. The stress injury measured as oxidative stress, electrolyte leakage, loss of chlorophyll and decrease in leaf water content was mitigated significantly in proline-treated plants. Additionally, proline application increased the level of sucrose and trehalose (cryoprotectants) in chilling-stressed plants. The studies revealed that proline application was significantly effective in reducing the impact of chilling injury on reproductive growth in chickpea.     

Effect of synthetic auxins on fruit size of five cultivars of Japanese plum (Prunus salicina Lindl.)

Most of the Japanese plum (Prunus salicina) cultivars grown in Israel produce relatively small fruit. Application of 2 l solution tree⁻¹ of 25 mg l⁻¹ 2,4-dichlorophenoxypropionic acid (2,4-DP) as butoxyethyl ester (Power™), 15 mg l⁻¹ 3,5,6-trichloro-2-pyridyloxyacetic acid (3,5,6-TPA) as free acid (Maxim^®), or 25 mg l⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) + 30 mg l⁻¹ naphthaleneacetic acid (NAA) (0.3% Amigo™) at the beginning of pit-hardening, when fruitlet diameter was ca. 22 mm, caused an appreciable and significant increase in fruit size. The yield of large fruit per cv.: ‘Kesselmen’ (100% increase), ‘Songold’ (100%), ‘Black Diamond’ (800%), ‘Royal Diamond’ (160%) and ‘Royal Zee’ (100%). As a result, the total yield of all five cultivars was also increased dramatically. Anatomical studies with ‘Songold’ revealed that the main effect of these synthetic auxins was via direct stimulation of fruit cell enlargement. The above auxins had no negative effect either on fruit quality at harvest (and after 1 week in shelf-life), or on return yield in the following year.     

Correlation of saponin content and Fusarium resistance in hybrids from different ploidy levels of Lilium Oriental

Lilium Oriental is of great commercial value, but large losses in production can result from its susceptibility to diseases caused by Fusarium ssp. Here we report a mutant of Lilium Oriental resistant to Fusarium, Cai-74, which was generated from crossing tetraploid (from ‘Star fighter’) and diploid (‘Cor. Amore’ × ‘Acapulco’) plants. Hybrid progeny were screened via inoculating Fusarium oxysporum into tissue cultured plantlets in a greenhouse. Although Cai-74 had a saponin content of 3.81 mg g⁻¹, which is much higher than its parents, the highly Fusarium-resistant wild species Lilium dauricum had the highest content of 4.11 mg g⁻¹ among the tested genotypes. Cai-74 had a 3A karyotype rather than 3B found in normal plants of Lilium Oriental. Taken together, our results suggest that Cai-74 bears a chromosomal variation in its structure, and that the saponin content in the lily scales correlate with the Fusarium resistance of Lilium Oriental. Cai-74 could be used as a gene resource for breeding Fusarium-resistant cultivars of Lilium Oriental.     

Plant regeneration of Carica papaya L. through somatic embryogenesis in response to light quality,gelling agent and phloridzin

Difficulties to develop an easy and reproducible protocol to get healthy and well formed plants from somatic embryos of papaya (Carica papaya L.) had included low germination, callus production at the base of the embryo radicle and the occurrence of hyperhydric plantlets among others, and by consequence unsuccessful transfer to the field. With the aim of improving a propagation method, the effects of light quality, gelling agent and phloridzin concentration on the germination of somatic embryos of hermaphrodite C. papaya L. var. Maradol were studied. Somatic embryos were grown on half strength MS medium, with the addition of Chen vitamins [Chen, M.H., Wang, P.J., Maeda, E., 1987. Somatic embryogenesis and plant regeneration in Carica papaya L. tissue culture derived from root explants. Plant Cell Rep. 6, 348–351], solidified with three distinct gelling agents: Sigma^® Agar–Agar, Difco^® Bacto agar and Phytagel^®; supplemented with phloridzin and exposed to different light qualities: blue (54 μmol m⁻² s⁻¹), red (65 μmol m⁻² s⁻¹), gro-lux (68 μmol m⁻² s⁻¹), red + blue, white (32 μmol m⁻² s⁻¹) and wide spectrum (49 μmol m⁻² s⁻¹) during a period of 4 weeks. Results show that light quality and gelling agent had important effects on germination and plant growth, while 3.0 mg L⁻¹ phloridzin had an important role on germination as well as in root development. Somatic embryos exposed to white light, culture medium solidified with 3.0 mg L⁻¹ phytagel and 3.0 mg L⁻¹ phloridzin showed longer roots. Meanwhile, germination and plant length were promoted on an improved culture medium solidified with 7.5 g L⁻¹ Difco^® Bacto agar, 3.0 mg L⁻¹ phloridzin and exposed to gro-lux lamps. Under these conditions, 70% of somatic embryos germinated and developed normal roots without hyperhydricity. The regenerated plantlets with well developed roots and shoots were successfully transferred to a greenhouse with a survival rate of 95%.     

Higher than optimum temperature under CO2 enrichment influences stomata anatomical characters in rose (Rosa hybrida)

There is little available information on the effects of temperature and CO₂ enrichment on stomata anatomical characteristics of plants. Effect of these two microclimates was studied on five rose (Rosa spp.) cultivars, viz. ‘First Red’ (used as check), ‘Arjun’, ‘Raktima’, ‘Raktagandha’ and ‘Pusa Pitamber’. Budded, single-stemmed rose cultivars having five lateral buds were grown in controlled environment growth cabinets under enriched CO₂ (1000 μmol mol⁻¹) and optimum (28/18 °C, T₀) or high (35/25 °C, T₁) temperature for 50 days. All observations were made on the abaxial leaf surface. Significant increases in stomatal density (68.7%), index (29.6%) and epidermal cell density (37.3%) were recorded in plants grown at high temperature over control with CO₂ enrichment. The cultivars responded differently in terms of length and width of guard cell and stoma (pore) under high temperature, however, the values averaged over treatments showed a significant reduction in these parameters. Further, number of stomata per leaf was higher (28.3%) in plants grown at high temperature, except First Red. A reduction in mean leaf area (26.7%) and dry mass (32.0%) was recorded at high rather than optimum temperature. The specific leaf area was maximum in Arjun (87%) while in First Red, a 14% reduction was noted at high temperature.     

Effect of 2,4-d, glutamine and BAP on embryogenic suspension culture of date palm (Phoenix dactylifera L.)

The proliferation of embryogenic suspension culture in two cultivars (Jihel and Bousthami Noir) of Phoenix dactylifera L. was tested on liquid media with or without 2,4-d and with different glutamine concentrations (3.35 × 10⁻⁴, 6.7 × 10⁻⁴ and 13.4 × 10⁻⁴ M). The liquid medium with 0.1 mg l⁻¹ 2,4-d and 6.7 × 10⁻⁴ M glutamine has clearly improved the proliferation of somatic embryos. In fact, when glutamine concentration increased from 3.35 × 10⁻⁴ to 6.7 × 10⁻⁴ M, the yield of somatic embryos increased from 14 to 56 embryos per 100 ml of culture medium for “Jihel” cultivar and 25–71 embryos per 100 ml of culture medium for “Bousthami Noir” cultivar. In contrast, increasing glutamine concentration from 6.7 × 10⁻⁴ to 13.4 × 10⁻⁴ M, the embryos yield was negligible. Based on biochemical analysis, the highest accumulation of proteins and sugars was obtained in liquid medium with 0.1 mg l⁻¹ 2,4-d and 6.7 × 10⁻⁴ M glutamine (118 and 91 mg of proteins g⁻¹ DW, respectively, for “Jihel” and “Bousthami Noir” cultivars; 194 mg of sugars g⁻¹ DW for “Jihel” cultivar and 182 mg of sugars g⁻¹ DW for “Bousthami Noir” cultivar). In addition, the supply of 0.05 mg l⁻¹ BAP on the germination medium could be useful in terms of germination percentage of somatic embryos. When BAP concentration increased from 0.05 to 0.2 mg l⁻¹, the germination percentage of somatic embryos decreased from 14.2 to 4.9%, while secondary embryogenesis increased from 26.4 to 45.2%.     

Molecular characterization and analysis of somaclonal variation in chrysanthemum cultivars using RAPD markers

Molecular characterization using RAPD analysis was carried out in eight cut flowers and two pot plant cultivars of chrysanthemum. Three of them (‘Refocus’, ‘Red Reagan’, and ‘Sheena Select’) were established in vitro and the occurrence of somaclonal variation was studied using the same molecular technique. Two induction media (MS + 0.1 mg l⁻¹ NAA + 0.1 mg l⁻¹ BA, and MS + 2.0 mg l⁻¹ IAA + 0.5 mg l⁻¹ Kinetin), and two proliferation media (MS + 0.1 mg l⁻¹ NAA + 0.2 mg l⁻¹ BA, and MS + 4.0 mg l⁻¹ IAA + 2.0 mg l⁻¹ Kinetin) were employed in order to evaluate the effect of the medium composition in the shoots’ stability. Likewise, the effect of the culture age was considered in assessing genetic stability. Monthly subcultures were carried out, identifying the origin and history of the shoots, throughout a nine-month proliferation period followed by acclimatization. Molecular markers were obtained in every subculture cycle and from the acclimatized plants. Only one shoot from the 7th subculture of the cultivar ‘Refocus’ showed a different band pattern. The use of RAPD for chrysanthemum cultivar characterization and somaclonal variation detection is discussed.     

Evaluation of ‘Hamlin’ sweet orange + ‘Montenegrina’ mandarin somatic hybrid for tolerance to Xanthomonas axonopodis pv. citri and Xylella fastidiosa

Asiatic citrus canker (ACC), caused by Xanthomonas axonopodis Starr & Garces pv. citri (Hasse) Vauterin et al., and citrus variegated chlorosis (CVC), caused by Xylella fastidiosa Wells et al., are considered the main diseases affecting sweet orange scion varieties in Brazil. Among commercial varieties, mandarins and tangerines are recognized as tolerant to these pathogens. We report herein the production of ‘Hamlin’ sweet orange (Citrus sinensis L. Osbeck) + ‘Montenegrina’ mandarin (Citrus deliciosa Ten.) allotetraploid somatic hybrid plants by protoplast fusion with improved disease tolerance that could be used as a donor of resistance genes in interploid hybridisation. Somatic hybridisation was confirmed by leaf morphology, flow cytometry and RAPD analyses. The somatic hybrid was propagated by grafting and cultivated in a screenhouse for tolerance assays. For X. axonopodis pv. citri assays, buds were collected from both ‘Hamlin’ sweet orange and the somatic hybrid and grafted onto ‘Cleopatra’ mandarin (Citrus reshni hort. ex Tanaka). As a negative control, buds from ‘Mexerica Tardia’ mandarin (C. deliciosa) were collected and grafted onto ‘Cleopatra’ mandarin. Two-month old plants with at least one young vegetative flush were individually spray-inoculated with a 10⁶ CFU mL⁻¹X. axonopodis pv. citri suspension and incubated in a growth chamber, at 27 °C, under 16-h photoperiod. The somatic hybrid showed a statistically significant reduction in susceptibility to ACC 30 days after inoculation. Compared to ‘Hamlin’ sweet orange, disease severity was reduced by 70%, with similar tolerance to that of the mandarin negative control. For X. fastidiosa assays, buds were collected from the somatic hybrid and its parental plants and grafted onto ‘Rangpur’ lime (Citrus limonia Osbeck). The developed plants were needle-inoculated with a X. fastidiosa suspension (8.7 × 10¹⁰ CFU mL⁻¹) into the new growth flush stem. Bacterial population was quantified both at 4 (at the inoculation point) and 8 months (50 cm above the inoculation point) after inoculation. The first evaluation detected X. fastidiosa in 63% of ‘Hamlin’ sweet orange and ‘Hamlin’ + ‘Montenegrina’ mandarin samples. In the second evaluation, X. fastidiosa was detected in 47.4% of ‘Hamlin’ sweet orange and 10.5% of ‘Hamlin’ + ‘Montenegrina’ somatic hybrid samples, suggesting that bacterial movement was restricted in the somatic hybrid. X. fastidiosa was not detected in both evaluations in samples collected from leaves of ‘Montenegrina’ mandarin. These results indicate that the ‘Hamlin’ sweet orange + ‘Montenegrina’ mandarin somatic hybrid has potential for improved disease tolerance that should enhance its value regarding future use in citrus breeding programs.