Effect of enhanced ultraviolet-B on allelopathic potential of Zanthoxylum bungeanum

The effect of enhanced ultraviolet-B on allelopathic potential of Zanthoxylum bungeanum was investigated. A significant inhibitory effect on germination rate of crop seeds under bioassay was observed at 25 g l⁻¹ and 50 g l⁻¹ by extracts from Zanthoxylum leaf both treated with enhanced ultraviolet-B radiation and untreated control. Medicago sativa and lettuce were more sensitive than radish to the extract from Zanthoxylum leaf treated with enhanced UV-B radiation, as the germination rates of M. sativa and lettuce were significantly reduced compared to control at 25 g l⁻¹ and 50 g l⁻¹, and so did alfalfa at 12.5 g l⁻¹. However, as for radish (Raphanus sativus) there was no significant reduction in germination rate at any concentration under bioassay compared to control. Content of UV-B absorbing compounds and total phenols in Zanthoxylum seedlings responded positively to enhanced UV-B radiation. The results suggest that the allelopathic potential of Z. bungeanum was generally improved under enhanced UV-B radiation.     

Production of inter-section hybrids between Primula filchnerae and P. sinensis through ovule culture

Inter-section hybrids were obtained in the reciprocal crosses between Primula filchnerae (2n = 2x = 24) of Sect. Pinnatae and P. sinensis ‘Fanfare’ (2n = 2x = 24) of Sect. Auganthus by rescuing ovules on half-strength (1/2) Murashige and Skoog's (MS) medium supplemented with 50 g l⁻¹ sucrose, 2.5 g l⁻¹ gellan gum, 0.1 mg l⁻¹ α-naphthaleneacetic acid (NAA), 0.1 mg l⁻¹ 6-benzyladenine (BA) and 50 mg l⁻¹ gibberellic acid (GA₃). In ovule culture, germination occurred with radicle elongation but no plumule was observed. The radicle kept on the initial medium showed root proliferation with callus formation. When the calluses were transferred to (1/2)MS media containing 30 g l⁻¹ sucrose and 3 g l⁻¹ gellan gum, without plant growth regulators (PGRs) or with 1 mg l⁻¹ zeatin and 0.1 mg l⁻¹ NAA, plantlets were regenerated. The plants thus obtained were confirmed to be hybrids through flow cytometry (FCM) and random amplified polymorphic DNA (RAPD) analyses. The hybrid obtained when P. filchnerae was used as the maternal parent was diploid, whereas hexaploid hybrid was obtained when using P. sinensis as the maternal parent. The hexaploid hybrid might be produced through chromosome doubling of a triploid originated from the fertilization of P. sinensis with unreduced pollen of P. filchnerae.     

The influence of desiccation and rehydration on the survival of polyembryonic seed of Citrus suhuiensis cv. limau madu

A study was carried out to investigate the influence of desiccation and freezing followed by various presowing rehydration procedures on the desiccation sensitivity of the seed of Citrus suhuiensis cv. limau madu. The freshly harvested seeds of limau madu were desiccated under a broad range of relative humidity (RH) to various equilibrium water contents (g H₂O g⁻¹ dw). The desiccated and desiccated–frozen seeds were either directly sown under germination conditions or subjected to presowing rehydration procedures: seed preheating, prehumidification and osmoconditioning. The hydrated and desiccated seeds were sown in controlled germination conditions and the survival was evaluated 4–6 weeks after sowing. The results showed that desiccation progressively reduced the percentage of normal seedling of the seeds of limau madu and the viability is almost lost at water contents below 0.08 g H₂O g⁻¹ dw. The estimated desiccation sensitivity was substantially high (WC₅₀ = 0.143 g H₂O g⁻¹ dw) when the desiccated seeds were rapidly rehydrated (uncontrolled rehydration). In contrast, seed prehumidification, preheating and osmoconditioning (controlled rehydration procedures) markedly enhanced normal seedling percentages decreasing the estimated values of WC₅₀ (between 0.08 and 0.127 g H₂O g⁻¹ dw). While the rapidly rehydrated desiccated–frozen seeds were almost killed at water content of 0.15 g H₂O g⁻¹ dw, prehumidification and preheating have noticeably increased percentage of frozen seeds survival at the same water content. However, at water content of 0.21 g H₂O g⁻¹ dw preheating significantly (P < 0.05) increased percentage of normal seedling of the frozen seeds. Seed desiccation markedly reduced the percentages of germinated seeds with multiple seedlings. Seed controlled rehydration remarkably increased the survival of polyembryos. The beneficial effect of seed controlled rehydration on the survival of the desiccated seeds was pronounced at medium water contents (0.08–0.25 g H₂O g⁻¹ dw).     

Ethylene influences development and flowering of Ptilotus spp. in vitro and ex vitro

The genus Ptilotus has immense potential for ornamental horticulture but its commercial development has been hindered by propagation limitations. Poor seed quality and germination are reported. Cutting propagation is limited by cutting supply as the juvenile phase of Ptilotus is short. Micropropagation has been used in an attempt to overcome these difficulties but explants become floral in vitro and this causes plantlets to elongate. Ethephon has been used to control flowering of stock plants of many ornamental species. This study investigated the effect of ethephon applied to young (3-week-old, deflasked from tissue culture) and mature (1-year-old) Ptilotus plants in a greenhouse. A system of applying gaseous ethylene at 0, 100, 200 and 300 mg l⁻¹ to the headspace of in vitro plantlets in glass jars was developed and the response of in vitro plantlets to ethylene studied. One-year-old Ptilotus plants were treated with 500 mg l⁻¹ ethephon 2 days before pruning or 1 or 2 weeks after pruning. Ethephon application 2 days before pruning decreased the number of inflorescences and increased the number of shoots (compared to the control) but was phytotoxic. Ethephon applications of 150 or 300 mg l⁻¹ applied weekly or fortnightly to 3-week-old plants deflasked from tissue culture reduced plant height and number of inflorescences and at low concentrations increased the number of new shoots. A fortnightly application at 150 mg l⁻¹ is recommended. Previous reports on the effects of ethylene on inflorescence production on plantlets in vitro are limited. Our study showed that exposure of in vitro plantlets of P. nobilis to ethylene gas at 100 mg l⁻¹ for 1 h significantly increased the number of shoots and plant height but this did not occur for plantlets of P. spicatus. Plantlets of P. spicatus exposed to transient ethylene at 200 and 300 mg l⁻¹ showed significantly greater rooting (52.4%) than the control (13.6%).     

Plant regeneration from stem segment-derived friable callus of “Fonio” (Digitaria exilis (L.) Stapf.)

“Fonio” (Digitaria exilis (L.) Stapf.) is a member of the grass family with excellent culinary and nutritional properties. In spite of its economic values, hardly has any improvement work been done. To enhance genetic improvement of this grain, plant regeneration protocol was developed using 8 cultivars. Stem segments of 5 mm long excised from 1 month-old seedlings germinated in vitro were cultured on 6 types of media for friable callus induction. Best result was obtained on Murashige and Skoog (MS) medium supplemented with 2 mg l⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) and 1 g l⁻¹ casamino acid, where 91.3, 88.9 and 87.8% of the explants formed friable calli in cultivars ‘Kurelep’, ‘Churiwe’ and ‘Agyong’, respectively. Shoots appeared when friable calli were transferred to two regeneration media, i.e., MSBZ (MS medium + 0.022 mg l⁻¹ 2,4-D, 0 .22 mg l⁻¹ 6-benzylaminopurine (BA), 0.22 mg l⁻¹ zeatin) and MSBG (MS medium + 0.5 mg l⁻¹ BA, 0.1 mg l⁻¹ gibberellic acid). The highest frequency of plant regeneration was attained on MSBG, with 91.7% of the friable calli forming shoots in cultivar “Churiwe”. Regenerated plants were rooted on hormone free MS medium. Flow cytometric analysis revealed 100% of the regenerants to be diploid. The protocol developed here can be used in the transformation of “Fonio” to increase the yield potential of this crop by incorporating characteristics such as disease resistance and stress resistance.     

Influence of yeast extract and casein hydrolysate on callus multiplication and somatic embryogenesis of date palm (Phoenix dactylifera L.)

Complex organic additives are known to improve growth and differentiation of in vitro plant cultures. The present investigation was conducted to determine the effect of various concentrations of yeast extract (YE) and casein hydrolysate (CH) on callus growth and somatic embryogenesis in date palm cultivar Nabout Saif. Callus induced from shoot tip explants was grown on callus multiplication medium supplemented with either YE or CH at 0.0, 0.1, 0.25, 0.5, and 1 g l⁻¹. To induce somatic embryogenesis, callus was transferred to a hormone-free medium containing the corresponding concentration of additives. The results have shown that callus weight and the number of somatic embryos were directly proportional to increases in the concentration of organic additives tested. Callus growth was best achieved when 1 g l⁻¹ of either YE or CH was added to the culture medium. At this concentration of YE, callus growth was double that of the control medium. On CH-containing media growth was 2.3 times that of the control. This indicates that CH is more effective in enhancing callus growth. However, YE was more effective in enhancing somatic embryogenesis. The data show that the best somatic embryo formation was obtained on either 1 g l⁻¹ YE or 0.5 g l⁻¹ CH which produced 45 and 30 embryos per culture, respectively, as compared to 20 embryos produced in the control treatment. Resultant somatic embryos successfully rooted and regenerated plantlets which exhibited normal growth in the greenhouse. Enhanced plant regeneration, an essential criterion for commercial micropropagation, was achieved.     

Factors affecting haploid induction through in vitro gynogenesis in summer squash (Cucurbita pepo L.)

Haploid production using in vitro ovule cultures has long been recognized as an important tool to produce haploid and homozygous double-haploid plants for genetic studies and plant breeding programs. In the present study, four experiments were carried out to study the influence of genotype, position of female flowers on plant stem, temperature and sucrose concentration on the in vitro gynogenesis induction of squash. (1) Ovules of 12 genotypes were excised from female flowers, 1 day before anthesis, and cultured onto MS medium containing 3% sucrose and 1 mg l⁻¹ from each of kinetin and 2,4-D (2,4- dichlorophenoxy acetic acid). Differences in response among genotypes were demonstrated. Raad F₁ showed the highest percentage of responding ovules and number of plantlets per dish with 48.8% and 15 plants, respectively. The results revealed that genotype is a key factor influencing the in vitro gynogenesis in squash. (2) Ovules were excised from first, second and third female flower of two hybrids (Giad and Raad) and cultured onto the mentioned above medium. The highest percentage of responding ovules and number of plantlets per dish were obtained from ovules excised from the second female flower on the plant stem. (3) Effect of temperature (4 and 32 °C) for 0, 4, 7 and 12 days on the ovule culture of Queen F₁ was studied. Ovules incubated at 4 or 32 °C for 4 days produced a better embryogenic response. (4) Three sucrose concentrations (30, 60 and 90 g l⁻¹) were tested with the ovule cultures of the local cultivar (Eskandrani). Differences among sucrose concentrations were statistically significant and ovules cultured on the MS medium containing 30 g l⁻¹ produced the best result. MS medium containing 90 g l⁻¹ did not produce gynogenic ovules.     

Mycorrhizal colonization improves onion (Allium cepa L.) yield and water use efficiency under water deficit condition

Most plants benefit from mycorrhizal symbiosis through improvement of water status and nutrient uptake. A factorial experiment with complete randomized blocks design was carried out in greenhouse at Tabriz University, Iran in 2005–2006. Experimental treatments were (a) irrigation interval (7, 9 and 11 days), (b) soil condition (sterile and non-sterile) and (c) arbuscular mycorrhizal fungi (AMF) species (Glomus versiforme, Glomus intraradices, Glomus etunicatum) and non-mycorrhizal (NM) plants as control. Onion (Allium cepa L. cv. Azar-shahr) seeds were sown in sterile nursery and inoculated with fungi species. One nursery left uninoculated as control. Nine weeks old seedlings then were transplanted to the pots. Average pre-irrigation soil water contents reached to about 67, 61.6 and 57.5% of FC corresponding to 7, 9 and 11 days irrigation intervals, respectively. At onion bulb maturity stage (192 days after transplanting), yield, water use efficiency (WUE) and yield response factor (K_y) were determined. The results indicated that AMF colonization increased soil water depletion significantly. G. versiforme under both soil conditions (sterile and non-sterile) and G. etunicatum in sterile soil depleted soil water effectively (P < 0.05). Mycorrhizal fungi improved WUE significantly (P < 0.0001) in both soil conditions. It raised by G. versiforme about 2.4-fold (0.289 g mm⁻¹) in comparison with the control (0.117 g mm⁻¹). G. intraradices and G. etunicatum also had significantly higher WUE than control. Apparently water deficit in 11-day irrigation interval led to lower yield and WUE compared to 9-day interval; the later resulted highest WUE (0.254 g mm⁻¹). Mycorrhizal plants increased seasonal ET significantly due to enhancing in plant growth; G. versiforme in both sterile and non-sterile soil and G. etunicatum in sterile soil had the highest ET. Bulb yield was influenced by irrigation period and fungi species. G. versiforme produced higher yield than other treatments (135.27 g/pot). Mycorrhizal plants in 11-day irrigation interval in spite of suffering from water stress had more bulb yield than non-mycorrhizal plants in all irrigation intervals. Yield in general was higher in 9-day treatments than other irrigating internals significantly (P < 0.05). Onion yield response factor (K_y) was decreased by AMF colonization; implying that symbiosed plants become less responsive to water deficit (longer irrigation interval) compared to the control ones.     

Growth and flowering of black iris (Iris nigricans Dinsm.) following treatment with plant growth regulators

The effects of application method and concentration of gibberellic acid (GA₃), paclobutrazol and chlormequat on black iris performance were assessed. Plants (10 cm high, 4 ± 1 leaves) were sprayed with 125, 250, 375 or 500 mg L⁻¹ or drenched with 0.25, 0.5, 1 or 2 mg L⁻¹ GA₃. In a second experiment, the plants were sprayed with 100, 250, 500 or 1000 mg L⁻¹ or drenched with 0.25, 0.5, 1 or 2 mg L⁻¹ paclobutrazol. Other plants were sprayed with 250, 500, 1000 or 1500 mg L⁻¹ or drenched with 100, 250, 375 or 550 mg L⁻¹ chlormequat. In each experiment, the control treatment consisted of untreated plants. Results indicated that the tallest plants (37.3 cm) in the GA₃ experiment were those sprayed with 250 mg L⁻¹. The most rapid flowering (160 days after planting) occurred when a 375 mg L⁻¹ GA₃ spray was used, whereas flowering was delayed to 200 days using 1 mg L⁻¹ GA₃ drench. Drenching with 1 mg L⁻¹ GA₃ increased height of the flower stalk by 7 cm compared to the control. Though relatively slow to flower, plants drenched with 1 mg L⁻¹ GA₃ had long and rigid stalks, which were suitable as cut flowers. Number and characteristics of the sprouts were not affected by GA₃. All paclobutrazol sprays resulted in leaf falcation. A 500 or 1000 mg L⁻¹ paclobutrazol spray resulted in severe and undesirable control of plant height, drastic reduction in stalk height and weight, and delayed flowering. Plants drenched with 0.25 or 1 mg L⁻¹ paclobutrazol were suitable as pot plants. Chlormequat reduced plant height only at the highest drench concentration, which also reduced flowering to 70%. No leaf falcation was observed with GA₃ or chlormequat. Chemical names: ( ± )-(R*,R*)-beta-((4-chlorophenyl)methyl)-alpha-(1,1,-dimethylethyl)-1H-1,2,4,-triazol-1-ethanol (paclobutrazol); (2-chloroethyl) trimethylammonium chloride (chlormequat).     

Effects of the commercial product based on Trichoderma harzianum on plant,bulb and yield characteristics of onion

Effects of the commercial product TrichoFlow WP™ (Agrimm Technologies Ltd., New Zealand), based on the fungus Trichoderma harzianum, on quality characteristics and yield of bulb onion was investigated. Bulb sets of the local cultivar Kantartopu was planted in soil with in and between row distances of 0.15 m and 0.40 m, respectively. The product, at considerably high dosages of 5 g m⁻², 10 g m⁻² and 15 g m⁻², was mixed with water and sprinkled once to the plots at planting. Analyses of data at harvest did not show statistical significance for Trichoderma effect on total bulb yield, bulb diameter, leaf length, number of shoot apex, %titratable acidity, number of internal (fleshy) leaves, number of external (papery) leaves, %soluble solids and %bulbs with diameters of 20–39 mm, 40–69 mm and ≥70 mm. The yields obtained from the plots treated with the dosages of 5 g m⁻², 10 g m⁻² and 15 g m⁻² and the control plots were 1063.7 kg da⁻¹, 1051.0 kg da⁻¹, 1066.5 kg da⁻¹ and 985.0 kg da⁻¹, respectively. Our results showed that high dosages of the Trichoderma product were not effective in enhancing onion bulb and yield characteristics under the given conditions.     

Direct regeneration of plants derived from in vitro cultured shoot tips and leaves of three Lysimachia species

The regenerability of three ornamental species—Lysimachia christinae, Lysimachia rubinervis and Lysimachia nummularia ‘Aurea’, were investigated using in vitro leaves and shoot tips. 6-Benzylaminopurine (BAP) and α-naphthalene acetic acid (NAA) added to Murashige and Skoog (MS) medium were tested for their effect on organogenesis. On the medium, shoot regeneration occurred directly without callus formation. In these species, L. christinae developed the highest regeneration rate and numbers of shoots/explant from shoot tips (100%, 12.25) and leaf bases (100%, 13.01) on the MS medium containing 3.0 mg l⁻¹ BAP and 0.1 mg l⁻¹ NAA. For L. rubinervis, the highest shoot induction rate and number of shoots/explant were obtained from shoot tip (100%, 16.87–17.20) on the MS medium with 0.1 mg l⁻¹ NAA and 3.0–5.0 mg l⁻¹ BAP. L. nummularia ‘Aurea’, however, showed the highest regeneration rate and number of shoots/explant (100%, 12.73) from leaf bases on MS medium supplemented with 1.0 mg l⁻¹ BAP and 0.1 mg l⁻¹ NAA. All in vitro shoots rooted well on half macronutrient MS medium containing 0.1 mg l⁻¹ NAA. After acclimatization, transplanted plantlets grew normally and flowered in the field.     

In vitro germination of walnut (Juglans regia L.) embryos

The present studies were undertaken with a view to standardize the medium and culture conditions for embryo culture of five cultivars of walnut viz., ACO 38853, Netar Akhrot, Gobind, Solding Selection and Blackmore. Embryos from mature fruits were aseptically excised and cultured on MS medium supplemented with different combinations of BAP, kinetin and GA₃. Best performing medium was MS with 0.5 mg l⁻¹ kinetin, 0.5 mg l⁻¹ BAP and 2 mg l⁻¹ GA₃ yielding 66.6% germination in Netar Akhrot after 12 days of culturing. Percent germination of excised embryos was higher when GA₃ and cold treatments were simultaneously applied as compared to those when applied separately. Netar Akhrot was found to be the best responding cultivar, which had a range of 25–66.6% embryo germination under different culture conditions. Plantlets with shoots and roots have been obtained in Netar Akhrot and ACO38853 and are transferred to soil after hardening.     

Development of a plant regeneration system from seed-derived calluses of centipedegrass [Eremochloa ophiuroides (Munro.) Hack]

The objective of this study is to establish plant regeneration system with the seed of the new Chinese selection “E-126”of centipedegrass [Eremochloa ophiuroides (Munro.) Hack] as explant. In present study, the following results were obtained: (1) The medium formulation most suitable for calluses induction was identified to be MS with 1.0 mg l⁻¹ 2,4-D + 30 g l⁻¹ mannitol + 50 ml l⁻¹ coconut milk and the ratio of calluses induction was 96.0%, including 5.2% of yellow granule calluses induction. The above medium formulation was adopted for subculture. (2) The rate of shoot regeneration from yellow granular calluses was 98.0% by MS optimum medium formulation with 2.0 mg l⁻¹ KT + 50 ml l⁻¹ coconut milk. The differentiated rate retained as high as 88.0% even after 5 times of subculture and 18.6% after 15 times of subculture. The optimum medium formulation for shoot growth was identified to be MS medium plus 2.0 mg l⁻¹ BAP, 0.8 mg l⁻¹ NAA and 50 ml l⁻¹ coconut milk. (3) The optimum medium for shoot rooting was identified to be MS medium with 0.6 mg l⁻¹ NAA + 50 ml l⁻¹ coconut milk, and the rooting rate to be 98.0%. The survival rate of transplanted plantlets from tubes to basin with soil was 92.0%. In conclusion, the plant regeneration system was successfully developed in this study, which may provide basic reference for screening of somaclonal variants and genetic transformation of centipedegrass.     

Seed germination and tissue culture of Cymbidium giganteum Wall. ex Lindl.

Efficient protocols were established for in vitro seed germination, neo-formation of secondary (2°) protocorms from primary (1°) protocorms and multiple shoot buds and protocorm-like body (PLB) induction from pseudo-stem segments of in vitro-raised seedlings of Cymbidium giganteum. Four nutrient media, namely Murashige and Skoog (MS), Phytamax (PM), Mitra et al. (M), and Knudson ‘C’ (KC) were evaluated for seed germination and early protocorm development. In addition, the effects of peptone, activated charcoal (AC) and two plant growth regulators [6-benzylaminopurine (BAP) and 2,4-dichlorophenoxyacetic acid (2,4-D)] were also studied. Both M and PM supplemented with 2.0 g l⁻¹ peptone or 1.0 mg l⁻¹ BAP resulted in ∼100% seed germination. Media supplemented with 2.0 g l⁻¹ AC could effectively induce large protocorms (1.6 ± 0.1 mm in diameter). Neo-formation of 2° protocorms from 1° protocorms was achieved in liquid and agar-solidified PM medium fortified with different concentrations and combinations of auxins (α-naphthalene acetic acid (NAA) and 2,4-D) and cytokinins [BAP and kinetin (KN)]. The highest number of 2° protocorms was obtained in liquid medium (10.7 ± 0.9/1° protocorm) supplemented with 2.0 mg l⁻¹ BAP + 1.0 mg l⁻¹ NAA. Although protocorms proliferated profusely in liquid medium, these did not develop further unless transferred to agar-solidified medium within 6–8 weeks. Multiple shoot buds and PLBs were induced from pseudo-stem segments on agar-solidified PM medium fortified with different concentrations and combinations of BAP and NAA and the maximum number of PLBs (6.00 ± 0.20) was recorded when BAP and NAA were applied at 2.0 mg l⁻¹ each. A solid root system was induced from PLBs and shoot buds when these were transferred to half-strength PM or M media fortified with 0.5 mg l⁻¹ indole-3-acetic acid. Well-rooted plants were transferred to the greenhouse with 95% survival.     

Effect of ABA and sucrose on germination of encapsulated somatic embryos of guava (Psidium guajava L.)

Present study demonstrates the effect of sucrose and ABA on germination of encapsulated somatic embryos of guava (Psidium guajava L.). Sucrose and ABA at different concentrations were also evaluated for their effects on maturation and germination of somatic embryos. Mature somatic embryos developed on MS medium containing high concentration of sucrose (10%) or ABA (1.0 mg l⁻¹) showed inhibition in germination if they continued to be in same medium for 4 weeks. With increasing concentrations of sucrose (3–9%) or ABA (0.01–1.0 mg l⁻¹) in medium, percent germination of encapsulated somatic embryos decreased significantly. Encapsulated somatic embryos after storage on MS medium supplemented with 9% sucrose or 1 mg l⁻¹ ABA for different duration (0–60 days) germinated when they were transferred to medium containing 3% sucrose. About 20.8% and 37.5% encapsulated somatic embryos germinated after storage on ABA (1 mg l⁻¹) or sucrose (9%) for 60 days, respectively. Temporarily suppression in germination of encapsulated somatic embryos by high concentration of sucrose or ABA may be important for short-term conservation of elite genotype of guava.     

A novel in vitro hydroponic culture system for potato (Solanum tuberosum L.) microtuber production

An innovative in vitro hydroponic culture system used in potato (Solanum tuberosum L.) microtuber production is described in this paper. In vitro potato plantlets, 6–8 cm in height, derived from meristems of potato tubers cultured on 1/2 Murashige and Skoog (MS) nutrient medium after 30 days culture were cut into 1.5 cm stem node segments and used as explants. These stem nodes were cultured in a novel system called in vitro hydroponic culture system containing 1/2 MS medium supplemented with 0.5 μM naphthaleneacetic acid (NAA), 0.3 μM gibberellic acid (GA₃), 3.7 μM adenine sulfate, 10% coconut water, 0.5 g/l activated charcoal, 80 g/l sucrose with or without 8 g l⁻¹ agar. Liquid medium was distributed to the carrier substrates in each storey of the system with the aid of capillary robes. In the present paper, the effects of porous material used as substrate carrier and the number of storeys involved in the culture system on microtuber formation and their morphological characteristics are reported. Cotton layer substrate is more stable for organogenesis of potato microtubers. Microtubers, 3.19 mm in diameter and 49.82 mg in weight, could be harvested from a one-storey in vitro hydroponic culture system containing filter paper as substrate. However, microtubers cropped from three-storey in vitro hydroponic culture system with cotton layer were bigger and weightier than those from three-storey system containing filter paper. The above results of the in vitro hydroponic system examined in this study might open up a new approach in producing potato and other hygrophilous microtuber.     

Chlorocholine chloride application effects on photosynthetic capacity and photoassimilates partitioning in potato (Solanum tuberosum L.)

Treatment of potato (Solanum tuberosum L.) with chlorocholine chloride (CCC) applied twice as a foliar spray 25 and 30 days after planting has shown to decrease shoot and stolon growth but increase tuber yield. However, the regulatory role of CCC on translocation of recently fixed photoassimilates into different parts of potato plants has not been fully illustrated. In this study, ¹⁴C-isotope labelling technique was used to estimate the photosynthetic capacity and photoassimilate partitioning among leaves, stems, roots + stolons, and tubers of potted potatoes treated with 1.5 g l⁻¹ CCC. CCC treatment significantly increased tuber dry mass but reduced leaf dry mass. CCC-treated leaves had significantly higher chlorophyll and carotenoid contents and assimilated 22.0% more ¹⁴CO₂ per leaf dry mass than the controls. Compared with the control, CCC treatment reduced the translocation of ¹⁴C-photoassimilates into leaves, stems and roots + stolons but increased that into tubers. CCC-treated leaves exported 14.6% more ¹⁴C-photoassimilates into other parts of the plants. In addition, CCC treatment reduced ¹⁴C-soluble sugar and ¹⁴C-starch accumulation in leaves and stems but enhanced them in tubers and roots + stolons. Collectively, the results indicate that CCC treatment significantly improves the photosynthetic capacity of potato leaves and promotes photoassimilates partitioning into tubers thereby enhancing tuber growth.     

Callus induction and plant regeneration from embryonic axes of Kosteletzkya virginica

Kosteletzkya virginica, a perennial dicot halophytic species of the Malvaceae, is native to American salt marsh. It was introduced into China as a potential species to improve coastal wetlands and to develop ecologically sound saline agriculture. K. virginica adapts excellently to the tidal-flat habitats in China's east coast, with multiple eco-benefits; in particular, its seed oil could be used to produce biodiesel. The purpose of this study was thus to develop a standardized protocol to induce a high frequency of callus and subsequent plantlet regeneration system for a K. virginica breeding program with the final objective of applying transgenic techniques to improve seed oil yield. The embryonic axes of K. virginica were used as explants for callus induction, shoot induction from the callus and then adventitious root induction from the shoots on nine culture media with different hormone combinations. The best results were achieved on the following media: (1) 93.94% callus induction on MS medium supplemented with 1.0 mg L⁻¹ indole-3-acetic acid (IAA), 0.3 mg L⁻¹ kinetin, 30 g L⁻¹ sucrose and 8 g L⁻¹ agar; (2) 65.83% shoot induction on 1/2MS medium supplemented with 0.1 mg L⁻¹ IAA, 0.5 mg L⁻¹ zeatin, 30 g L⁻¹ sucrose and 8 g L⁻¹ agar; (3) 96.67% rooting on MS medium containing 30 g L⁻¹ sucrose and 8 g L⁻¹ agar. The survival rate of plantlets by organogenic regeneration was 85% after being transplanted into potting soil in flowerpots and placed in the greenhouse. This experiment indicates that we established successful callus induction and plant regeneration protocols for K. virginica.     

In vitro rescue of immature avocado (Persea americana Mill.) embryos

An in vitro culture protocol was developed as a means of avocado embryo rescue. Different factors including presence of cotyledons, medium texture and cold or gibberellic acid pretreatments, were studied. To better understand the germination process in this recalcitrant species, immature zygotic embryos at different stages were used in these experiments. Optimum results were dependant on the embryo developmental stage. Whereas smaller embryos (5 mm long) germinated better in M1 liquid medium, 15 mm long embryos responded better when precultured in B5m medium supplemented with 1 mg l⁻¹ GA₃, and fully mature embryos were capable of germinating directly in solid M1 medium. Our results suggest the existence of two types of dormancy in avocado embryos: an embryo-dormancy caused by cotyledons, and another type of dormancy, mainly occurring in 35 mm long embryos and revealed by the formation of dwarfing rosette seedlings, that can be released by a GA₃ pretreatment.     

Foliar application of chlorocholine chloride improves leaf mineral nutrition,antioxidant enzyme activity,and tuber yield of potato (Solanum tuberosum L.)

In southern China, potatoes (Solanum tuberosum L.) are grown in the late season and the crops are often subjected to low temperature stress particularly during the tuber bulking stage. Exogenous chlorocholine chloride (CCC) treatment has been found to improve crop performance under suboptimal growth conditions; however, the physiological mechanisms underlying the beneficial effects have not been fully understood. The objective of this study was to investigate the effects of CCC treatment on mineral nutrition, antioxidant enzyme system, and tuber yield of potato (cv. Zhongshu 3) under field conditions. The plants were foliar sprayed twice with 1.5, 2.0 and 2.5 g l⁻¹ CCC at 24 and 28 days after emergence (DAE), respectively; and plants without CCC treatment were serviced as control. Leaf samples were collected on 56 DAE for determination of mineral nutrition contents and antioxidant enzyme activity. Results showed that 1.5 and 2.0 g l⁻¹ CCC treatments significantly increased the contents of P, K, Ca, Mg, Fe, Mn, Zn and Cu in potato leaves. These treatments also increased superoxide dismutase (SOD), peroxidases (POD) and catalase (CAT) activities in the leaves. A positive linear relationship was found between SOD activity and the content of Fe + Mn + Zn + Cu. Tuber yield was significantly increased by CCC treatment. It is concluded that treatment with certain concentration of CCC (e.g. 1.5–2.0 g l⁻¹) improves mineral nutrition and SOD, POD and CAT activities in potato leaves; which might have contributed to the higher tuber yield of the crop grown under suboptimal conditions.