不同培养条件对‘丰香’草莓离体叶片再生的影响

 以草莓品种‘丰香’离体叶片为外植体, 探讨了基本培养基、不同细胞分裂素、暗培养、硝酸银浓度以及不同植物生长调节剂组合对不定芽再生的影响。结果表明, 基本培养基中以MS 最为适合,WPM、QL 、AS 培养基均不利于不定芽的再生, 而TDZ 的诱导效果好于BA。以MS 基本培养基附加TDZ2.0 mg·L - 1和IBA 0.8 mg·L ^-1可以使‘丰香’叶片不定芽的再生率高达72.33 % , 平均每叶再生芽5.59个。暗培养14 d 可以将‘丰香’叶片的不定芽再生率提高到90.09 %。硝酸银对于提高‘丰香’叶片的不定芽再生没有明显效果, 但在一定程度上改变了细胞分化的方向。     

De novo differentiation of shoot buds from leaf-callus of Dianthus caryophyllus L. and control of hyperhydricity

A method is described for producing de novo shoots from leaf derived callus of carnation (Dianthus caryophyllus L.). Plants were regenerated in four steps, viz. callus induction, shoot regeneration, removal of hyperhydricity from regenerated shoots and root development. Callus induction medium contained 2,4-D and BAP. Shoot buds were formed when the callus was further subcultured on 2,4-D- and BAP-containing medium, or MS medium without any growth regulators. The shoots so formed were hyperhydric, bushy in appearance with reduced stem length and watery leaves. The normal conformation of shoots was restored by culturing the hyperhydric shoots onto medium supplemented with GA₃ and bactopeptone. The recovered shoots were rooted on MS medium added with NAA (1 mg/l) or IBA (2 mg/l). Regenerated plants with well-developed root and shoot systems were successfully transferred to field conditions after initial acclimation.     

Dissemination of Phytophthora cactorum, cause of crown rot in strawberry,in open and closed soilless growing systems and the potential for control using slow sand filtration

Closed (recirculating) growing systems provide a greater potential for the dispersal of water-borne plant pathogens and disease expression compared to open (run-to-waste) systems. Here we studied the effects of three soilless growing systems (open, closed, and closed with slow sand filtration) on the dispersion of Phytophthora cactorum propagules and the severity of the crown rot disease in strawberry (Fragaria × ananassa Duch.). The plant-growth medium used was coir fiber. The three growing systems showed the same density of P. cactorum propagules in the water drained from the growing media. However, propagules of this pathogen were not detected by the baits in the filtered solution recovered from slow sand filtration. In all systems Phytophthora propagules dispersed from the inoculated plant to adjacent uninoculated plants. At the end of the first crop no differences in the severity of crown rot were found between the different systems of crop culture. However, at the end of the second crop cycle, crown rot in the closed soilless system without slow sand filtration was more severe than in the other two systems. These results demonstrated that the commercial potential of slow sand filtration to prevent propagule dispersal and hence suppress crown rot in strawberry crops grown in a closed culture system.     

缺钾胁迫对连作草莓生长和土传病害的影响

 以草莓‘红颜’品种为试材,设缺K（K+ 48.75 mg · L^-1,缺量水平）、1/2K（K+ 415.58 mg · L^-1, 半量水平）和全量K（K+ 782.41 mg · L^-1,正常水平）营养液处理,对缺钾胁迫与草莓连作障碍的关系进 行了探讨。结果表明,缺钾处理显著抑制二茬草莓植株地下部分和地上部分的生长,其中缺K 的长势最 弱,1/2K 比全量K 的长势强;缺K 二茬植株根系分泌物对草莓组培苗生长有显著影响,表现为低浓度（1%、 2%）根系分泌物促进幼苗侧根和茎叶生长,高浓度（4%）则抑制根系生长。不同浓度钾营养液处理的三 茬植株分别接种尖孢镰刀菌和大丽轮枝菌后,在不同时期比未经过连作的基质对照发病都严重。缺K 三 茬植株发病最重,接菌后10 d 时病情指数就高达75,全量K 三茬植株发病次之,1/2K 三茬植株发病相 对较轻。因此,缺K 环境会加重草莓的连作障碍,适宜的K 量对草莓连作障碍的防治有重要意义。     

Continuous propagation of tomato plants by means of callus cultures

Callus obtained in vitro from stem internode tissue was used to investigate the possibilities for accelerated vegetative propagation of Lycopersicum esculentum Mill. and Lycopersicum peruvianum (L). The results of different tests with phytohormones pointed to a high endogenous auxin level in the original stem explants of L. peruvianum. In L. esculentum shoots were obtained when high amounts of zeatin or coconut-milk were applied.Explants of CCC-pretreated (2 chloroethyl trimethyl-ammonium chloride) tomato plantlets showed a significant increase in shoot formation as compared to the untreated ones.Callus tissue that was more than 2 years old and had been used in 30 transfers still had the capacity to produce normal shoots and roots.More than 200 resulting plants were observed in glasshouse conditions for possible genetic variations. No striking deviations from the original phenotype occurred. Seeds harvested from the fruits of self-fertilized flowers on these plants were sown under normal growth conditions. Some plants of one particular cultivar showed signs of accentuated vegetative development.     

Chilling response of ‘Granny Smith’ apple lateral buds inhibited by distal shoot tissues

Apple (Malus×domestica Borkh.) shoots were decapitated before or after chilling and then forced to budburst to determine the influence of distal inhibition (paradormancy) on the chilling response of lateral buds. Shoots were chilled and forced with the presence or absence of an inhibitory distal disbudded shoot piece. Endogenous cytokinins were determined from distal and proximal segments of shoots segmented before and after chilling and/or forcing. The isolation of lateral buds from distal inhibition by decapitation before chilling resulted in a dramatic increase in growth rate. This increase is greater than that observed in terminal buds during the normal development of acrotony on intact shoots. Distal shoot tissues appear to inhibit the chilling response of lateral buds. On intact shoots cytokinin increased more in the distal shoot tissues, but in decapitated shoots cytokinin increased more in the proximal shoot tissues. Increased cytokinin in the proximal stem segment following decapitation is possibly associated with an increased lateral bud growth rate.     

Cytokinins in citrus. II. Fluctuations during growth in juvenile and adult plants

Juvenile and adult plants of ‘Pickstone Valencia’ orange (Citrus sinensis (L.) Osbeck) were compared for differences in shoot and root growth and for endogenous cytokinins in a synchronized growth flush. Dry-mass accumulation in both shoots and roots of juvenile-budded plants was significantly greater than in adult plants. Cytokinins extracted from leaves and roots of plants removed from the dormancy-induction environment, and subsequently during active growth, were mostly of the non-polar type. Buds of juvenile scions appeared to accumulate higher levels of polar cytokinins during dormancy than adult scion buds. In both plant categories, the level of polar cytokinins decreased once growth was stimulated by higher temperatures. However, cytokinin levels in buds were markedly higher than in new shoot, leaf or root material. Polar cytokinins were present in particularly low concentrations in the developing new shoots, and non-polar cytokinins only increased after the extent of shoot enlargement reached a maximum at about 50 days after warm-temperature incubation.The possible involvement of cytokinins in growth differences between adult and juvenile plants seems to be at the stage of bud activation, prior to breaking of dormancy. At this stage, polar cytokinins reached a much higher level of activity in juvenile buds, and this may have resulted in the greater scion growth of juvenile plants.     

Bulb-dip application of growth-regulating chemicals for inhibiting stem elongation of ‘Enchantment’ and ‘Harmony’ lilies

An improved method of treating Lilium cultivars ‘Enchantment’ and ‘Harmony’ with the growth regulators Ancymidol, CCC, and Ethrel is described. When bulbs were treated, prior to planting, by immersing them for 12 h in aqueous solutions of the growth regulators, shoot elongation was more effectively inhibited than when the chemicals were applied as soil drenches. Ancymidol was the most potent inhibitor of stem elongation. Although Ethrel effectively inhibited stem growth during the first treatment season, it resulted in increased elongation of the renewal shoots. Ethrel interfered with apical meristematic activity, and induced early flower senescence. The inhibitory effects of a single bulb-dip with Ancymidol and CCC were evident on the growth of the renewal shoots in the following non-treatment season.The bulb-dip procedure offers several advantages: chemicals can be applied effectively at a much earlier development stage of the shoot; more effective control of shoot elongation is possible; significant control of shoot elongation may be obtained with lower concentrations of a growth regulator; one application of the growth regulator affected growth of the current season's shoot and also the growth of the renewal shoot in the following season; bulbs may be pretreated for use either as pot plants or for landscaping; chemically pretreated bulbs can be readily packaged and distributed without further chemical treatments being required.     

Auxin-dependent development and habituation of highbush blueberry (Vaccinium × covilleanum But. et Pl.) ‘Herbert’ in vitro shoot cultures

The influence of IAA (1.0 mg dm⁻³), and IBA (1.16 mg dm⁻³), on the development of highbush blueberry (Vaccinium × covilleanum But. et Pl.) ‘Herbert’ in vitro shoot cultures was examined. Depending on the kind of auxin and 2iP concentration in vitro cultures consisted of various number of axillary (AX) and adventitious (AD) shoots. Three different categories of AD shoots were found: leaf shoots (AD-L), node-adjoin shoots (AD-P), and base-adjoin shoots (AD-M, madshoots). The AX shoots were the least habituated (towards auxins, cytokinins and vitamins) whereas the AD-M shoots (madshoots) the most. In comparison to IAA, IBA caused dying or callusing higher number of initial explants. However, IBA generally suppressed development of AD shoots, especially madshoots whereas slightly weakened multiplication of AX shoots. IBA significantly enhanced elongation of AX shoots also. Axillary shoots obtained on IBA-media had relative long internodes and rigid, well-developed leaves. The adventitious shoots, especially base-adjoin (AD-M) ones, were easily distinguishable as were more thin and fragile, more or less vitrified, and had short internodes and smaller, sometimes unfolded leaves. Development of blueberry in vitro cultures on auxin-free and IAA-supplemented media was similar. AX shoots grown on such media resembled AD shoots. 2iP applied in higher doses along with IAA promoted much proliferation of AD than AX shoots. In contrast, 2iP applied in higher doses together with IBA stimulated significantly only growth of AX shoots whereas in general, development of adventitious shoots was not affected. Micropropagation carried out through routine method based on subculturing of shoot explants or shoot clumps on the medium supplemented with IAA (4 mg dm⁻³) and 2iP (10–15 mg dm⁻³) as well as stimulation of shoot elongation on the blank medium causes in fact the propagation of highbush blueberry through highly habituated adventitious madshoots. Replacement of IAA by IBA facilitates micropropagation of highbush blueberry cv. Herbert through axillary shoots.     

Regeneration and characterization of plants derived from leaf in vitro culture of two sweet orange (Citrus sinensis (L.) Osbeck) cultivars

In the current work attempts were made to investigate culture of leaf explants derived from in vitro seedlings of two sweet orange (Citrus sinensis (L.) Osbeck) cultivars, Bingtangcheng and Valencia. Effects of several factors, including culture medium, lighting condition, explant age and genotype on regeneration response were examined based on three parameters, percentage of explants producing shoots, mean number of shoots per explant and shoot forming capacity. Culture of the explants on shoot-inducing media (SIM) composed of MT salts supplemented with different growth regulators gave rise to disparate shoot regeneration, in which SIM₁ (MT + 0.5 mg L⁻¹ BA + 0.5 mg L⁻¹ Kinetin + 0.1 mg L⁻¹ NAA + 3% sucrose + 0.8% agar, pH 5.8) was shown to be the most effective medium for direct induction of shoots from leaf explants. Highly significant difference in the response of shoot bud regeneration was noted between the two cultivars, with Bingtangcheng being more responsive than Valencia. Culture of explants from fully developed leaves led to better shoot regeneration capacity in comparison to undeveloped ones. However, the two lighting conditions used herein did not cause significant difference in shoot regeneration. Phenotypic observation and randomly amplified polymorphic DNA (RAPD) analysis confirmed that all the regenerated plants from both genotypes were genetically identical to their donor plants, suggesting absence of detectable genetic variation in the regenerated plants. The data presented here demonstrated that direct initiation of plants from leaf explants has been successfully accomplished. To our knowledge, this is the first report on direct regeneration of shoots from leaf explants in Citrus, which will provide an alternative source for citrus genetic manipulation in the future.     

Nitrogen and calcium nutrition of Petunia × hybrida ‘Coral Sea’

The effects of N and Ca nutrition on plant growth and shoot elemental content of Petunia × hybrida Hort. Vilm. - Andr. ‘Coral Sea’ were evaluated. Nitrogen and Ca were applied separately or in combination in three experiments: (1) N at 0, 100, 200 or 400 mg l^?1; (2) Ca at 0, 75, 150 or 300 mg l^?1; (3) N at 0 or 100 mg l^?1 and Ca at 0 or 150 mg l^?1 combined factorially. Shoot and root dry weights, branch length and flower number were highest when plants received 100 mg l^?1 N. Plants treated with 150 mg l^?1 Ca had the highest shoot and root dry weights. Branch length was maximal at 300 mg l^?1 Ca.Nitrogen and Ca interacted to increase shoot dry weights, branch number and length, leaf area and flower number. Increasing N concentrations increased N and decreased P, Mn and Zn shoot contents. Calcium content of shoots increased while N, P and Mg decreased in response to increasing applications of Ca to petunia plants. Minimal N and Ca tissue concentrations for optimal P. × hybrida growth were 3.3 and 0.67%, respectively.     

Chemical “pinching” of pot chrysanthemums with 2,3-dihydro-5,6-diphenyl-1,4-oxathiin

In summer and winter experiments a research formulation of 2,3-dihydro-5,6-diphenyl-1,4-oxathiin (code name UBI-P293) was effective as a chemical “pinching”-agent on pot chrysanthemum cultivar ‘Bright Golden Anne’, when applied at 0.4 and 0.8% active ingredient to the apical region of plants within a few days of planting. A lower concentration was not so effective. When the applications were made later, they induced considerable variation in the length and quality of lateral shoots and also delayed flowering. The compound did not always prevent the development of the terminal bud, but reduced apical dominance sufficiently to permit the lateral shoots to develop normally. When P293 successfully “pinched” the plants in the winter experiment, it had no effect on the length of the lateral shoots which developed. At flowering these were too long from a commercial grower's point of view. In the summer experiment shoot length was adequately reduced by application of a foliar spray of a new quaternary ammonium growth retardant when the lateral shoots were a few centimetres long.     

Improvement of caper (Capparis spinosa L.) propagation using in vitro culture and gamma irradiation

Some of the factors influencing the propagation of caper (Capparis spinosa L.) plants in vitro and germination of the seed were studied. The number of adventitious shoots emerging from caper stems cultured in vitro increased from 2.2 shoots per explant when the growth medium contained 2 mg/L of gibberellic acid (GA3) to 5.5 when the growth medium contained 2 mg/L zeatin riboside (ZR) and 1 mg/L naphthalene acetic acid (NAA). The best medium for callus formation from leaf and stem parts contained the growth regulators 1 mg/L 6-benzylaminopurine (BAP) and 0.1 mg/L NAA and the best medium for plant regeneration contained 1 mg/L kinetin and 0.1 mg/L indole-3-acetic acid (IAA). The effect of gamma irradiation on the growth of caper shoots in vitro was also studied. A 10 Gy dose of gamma irradiation stimulated growth of shoots up to 200% and increased shoot rooting percentage from 75 to 100%.     

Direct and callus-mediated regeneration of Curcuma soloensis Valeton (Zingiberaceae) and ex vitro performance of regenerated plants

Protocols are outlined for the regeneration of Curcuma soloensis, an attractive tropical ornamental plant, from young vegetative bud explants. We used both direct and callus-mediated regeneration techniques to produce material suitable for mass propagation and the development of transgenic plants. During direct plantlet propagation, the presence of thidiazuron (TDZ) in the growing medium induced more than three times as many shoots as 6-benzylaminopurine (BA), with a mean of 18.7 shoots per explant on MS medium containing 2.5 μM TDZ compared to 5.0 shoots with 40 μM BA. Subsequently, the shoots rooted readily on MS basal medium that was free of plant growth regulators. During indirect plantlet regeneration, TDZ combined with BA and 2,4-dichlorophenoxyacetic acid (2,4-D) had significant effects on embryogenic callus induction and multiplication. The frequency of callus formation was 91.1% for explants cultured on MS basal medium supplemented with 2.5 μM TDZ, 2.0 μM BA and 1.2 μM 2,4-D. On average 7.1 shoots were produced per callus mass cultured on MS medium supplemented with 2.5 μM TDZ, 9.0 μM BA and 1.2 μM naphthaleneacetic acid (NAA). Regenerated shoots were transferred to MS medium supplemented with 2.5 μM TDZ, to produce multiple shoots. In vitro cultured plantlets readily acclimatized to greenhouse conditions, showing 100% survival rates in a sphagnum, perlite and sand (1:1:1) medium. These plants were transplanted into pots or planted in the field. The ex vitro acclimated plants grew vigorously and produced showy inflorescences 5–6 months after planting. The high-frequency of shoot multiplication and rapid flowering of tissue-cultured plants indicate that C. soloensis has great potential in the floricultural market.     

In vitro micropropagation of the ornamental prickly pear cactus Opuntia lanigera Salm–Dyck and effects of sprayed GA3 after transplantation to ex vitro conditions

We established the conditions to micropropagate the ornamental prickly pear cactus Opuntia lanigera Salm–Dyck through axillary shoot development from isolated areoles. For the shoot proliferation stage different explant orientation (vertical and horizontal), type of cytokinin (BA, DAP and K), and concentrations (0, 1.25, 2,5, 5.0 and 7.5 mg/L) were evaluated. Media [Murashige, T., Skoog, F., 1962. A revised medium for rapid growth and bioassays with tobacco cultures. Phys. Plant. 15, 473–497: 50 and 100%], and carbohydrate concentration (0.25, 0.5, 0.75 and 1.0%) were studied to optimize individual shoot growth and elongation. Following micropropagation and plantlet acclimatization, the effects of GA₃ on plant growth were determined by spraying a series of increasing concentrations (0, 150, 300 and 450 ppm). A reliable and efficient protocol of micropropagation was established for this particular plant species. The greatest propagation ratio (shoot proliferation) was obtained when explants were cultured in vertical orientation (4.975 shoots per explant) as compared to horizontal position (3.692 shoots per explant). The addition of BA to the media resulted in increased shoot number per explant (8) in comparison to K and DA, which produced only 2 shoots in average. However, after 42 days of culture, significantly higher shoot length was obtained with DAP (14 mm) compared to K and BA (4 mm). After the shoot proliferation stage, an elongation subculture was performed prior rooting in which shoot growth was enhanced when crowns of shoots were cultured in 50% of basal salt formulation of Murashige and Skoog (1962) and low sucrose concentration (2.5 and 5%). Exogenous application of GA₃ after plantlet acclimatization on glasshouse conditions increased spine-hair (developed from areoles in young plants) length as part of short-term effects. However, significantly higher values were obtained in plantlets treated with 300 ppm of GA₃ when compared with the rest of the treatments. At the end of the study, the most important long-term effect produced by GA₃ was the suppression of total shoot growth. The micropropagation protocol described here and the conditions to grow the plants through fertigation plus the application of GA₃ that induced changes in the phenotype may be used in commercial exploitations to regenerate 12,500 plantelts in average after 12 months of culture and produce healthy plants with better ornamental characteristics and higher commercial value.     

Commercial production of Cordyline terminalis (L) Kunth. from shoot apex meristem and assessment for genetic stability of somaclones by isozyme markers

Rapid multiplication of Cordyline terminalis (L) Kunth. was achieved from shoot apex explant on Murashige and Skoog's (MS) medium supplemented with 3% (w/v) sucrose and different concentration of growth regulators at various combinations. On MS medium supplemented with BA in combination with Adenine sulphate and IAA, shoot initiation and multiplication were obtained. Best elongations of shoots were found on 1/2 MS basal medium and shoots were rooted on 1/2 MS medium supplemented with IBA. Rooted plants passed through a hardening phase prior to ex vitro transfer. Clonal propagated plants established in garden soil were uniform and identical to mother plant in respect to growth characteristics and morphology. Isozymic profiles of different micropropagated clones were assessed for their genetic stability. Ten clones were tested for six isozymes. Only a few showed variation with respect to the banding pattern in esterase and superoxide dismutase. In superoxide dismutase, the two polymorphic isoforms (Rf 0.06 and 0.45) appeared in the clone C8 of the plants transferred to the field after 15 subcultural passages. Mobility and intensity of bands were monitored in other isozymes. Isozyme markers may be used as a tool for rapid screening of genetic stability in tissue cultured clones of C. terminalis.     

Morphological and physiological characteristics of micropropagated strawberry plants rooted in vitro or ex vitro

In the classical method of strawberry micropropagation, the rooting phase is done in vitro. The trials were undertaken to replace in vitro rhizogenesis by a direct ex vitro rooting. The aim of the present study was to evaluate the morphological and physiological status of strawberry plants rooted by both methods. The micropropagated shoots of strawberry, cultivars Senga Sengana, Kent and Kama were rooted by the standard method at the in vitro stage or they were treated as soft cuttings and rooted ex vitro (in non-sterile conditions). After a 4-week rooting period the plantlets rooted ex vitro had larger root systems than in vitro-rooted ones, as evidenced by a significantly lower ratio of shoot to root dry weights (2.95, 3.33 and 4.24, respectively, for ex vitro rooted Senga Sengana, Kent and Kama plants versus 5.09, 6.95 and 5.04 for in vitro rooted plants of the same cultivars). During subsequent growth, differences in development increased and were most pronounced in runner formation, more than twice as many runners were formed by ex vitro, than by in vitro-rooted plants. Chlorophyll fluorescence parameters showed that photochemical activity was similar in the leaves from plants rooted in vitro and ex vitro. Values of chlorophyll fluorescence parameters indicated that persistent leaves play the key photosynthetic role at the beginning of the growth period. With the formation of new leaves, the photosynthetic activity of the persistent leaves decreased and their function was taken over by the newly formed ones.     

Biological control of the shoot disease of Amaranthus hybridus caused by Choanephora cucurbitarum with Bacillus subtilis

Summary

Isolates of Bacillus subtilis from soil and ogili (a local food condiment) controlled choanephora shoot disease of the vegetable crop plant, Amaranthus hybridus, in the greenhouse. Disease developed on plants inoculated simultaneously with the pathogen Choanephora cucurbitarum and the ‘ogili’ isolate, but not the soil isolate, of B. subtilis. Application of either bacterial strain to plants one day before a challenge with the pathogen prevented disease development. A single application of either strain prevented disease development from a subsequent challenge with the pathogen at any time over a thirty day period. Viable counts of the microflora on the shoot tips of the treated plants indicated strongly that the inoculated bacteria multiplied and colonized the extending shoot tip whether or not the shoots were covered with polythene bags. Both strains of Bacillus subtilis inhibited mycelial extension growth as well as conidial germination in vitro.     

Grafting and calcium cyanamide as alternatives to methyl bromide for greenhouse eggplant production

Eggplant (Solanum melongena L.) seedlings (cv. Tsakoniki) were cultivated in soil artificially infested with V. dahliae Kleb. and then sterilized by either methyl bromide (MB) or calcium cyanamide. Grafted seedlings on the wild species Solanum torvum Sw. and the control seedlings (auto-rooted) were cultivated in soil sterilized by MB and then artificially infested with V. dahliae. The plants grafted on S. torvum and the ones grown in soil treated with calcium cyanamide (2001) exhibited significantly lower leaf symptom index (average value LSI = 1.55 and LSI = 1.00) and disease index (average value DI = 2.05 and DI = 1.20), respectively, as compared to the controls (average value LSI = 3.80 and DI = 5.50). Grafted plants on S. torvum and plants grown in soil treated with calcium cyanamide (2001) were more vigorous, as measured by plant height, stem diameter and root biomass than the controls. This resulted in an increased (over years) early (487.8% and 416.2%, 2001) and late production (277.0% and 241.3%, 2001) and mean fruit weight (over years) in early (93.7% and 49.6%, 2001) and late production (38.3% and 22.8%, 2001) as compared to the controls. In conclusion, grafting of eggplant and soil sterilized by calcium cyanamide had positive effects on growth, production and Verticillium wilt control.