Effect of synthetic auxins on fruit development of ‘Bing’ cherry (Prunus avium L.)

The main cherry cultivar grown in the warm climate of Israel, ‘Bing’, produces relatively small fruit. Over three consecutive years (2003–2005), application of 50 mg l⁻¹ 2,4-dichlorophenoxypropionic acid [2,4-DP; as its butoxyethyl ester (Power™)], 10 mg l⁻¹ 3,5,6-trichloro-2-pyridyloxyacetic acid [3,5,6-TPA; as the free acid (Maxim^®)], or 25 mg l⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) plus 30 mg l⁻¹ naphthaleneacetic acid (NAA; 0.3% Amigo™), at the beginning of pit-hardening when fruitlet diameter was ca. 13 mm caused appreciable and significant increases in fruit size and total yield, except when the crop load was heavy. Anatomical studies revealed that the main effect of these synthetic auxins was via direct stimulation of fruit cell enlargement. The above auxins had no negative effect on fruit quality, either at harvest or after 1 month of storage at 0 °C, or on return yield in the following year.     

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.     

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.     

Micropropagation of the bromeliad Guzmania ‘Hilda’ via organogenesis and the effect of α-naphthaleneacetic acid on plantlet elongation

This study established a highly effective micropropagation system to obtain good plantlet proliferation from floral organs via callus induction and bud differentiation in Guzmania ‘Hilda’ bromeliad. The best frequencies of organogenic callus formation (20% in petal and 35% in ovary explants) were obtained on media containing a combination of 1.0 mg l⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) with 1.0 mg l⁻¹ α-naphthaleneacetic acid (NAA) and 1.5 mg l⁻¹ 2,4-D with 0.5 mg l⁻¹ NAA, respectively. Organogenic calli were cultured on medium with 1.0 mg l⁻¹ NAA and 0.5 mg l⁻¹ 1-phenyl-3-(1,2,3-thiadiazol-5-yl) urea (TDZ) induce the differentiation and regeneration of adventitious buds into plantlets. When the plantlets were cultured in a medium with optimum NAA concentration (0.5–1.0 mg l⁻¹) significant improvement in regeneration and elongation was achieved within one month. This overcame the difficulty of delayed elongation in Guzmania plantlets. More than 99% of the regenerated and acclimatized plantlets developed to the flowering stage.     

Potential of ethephon,NAA, NAD and urea for thinning pistachio fruitlets

Alternate bearing, the occurrence of high yield ‘on’ year followed by low yield ‘off’ year, is striking in pistachio (Pistacia vera L.). Floral buds of pistachio are formed a year before bloom, but abscise during the years with heavy crop (‘on’ year). Abscission of floral buds is due to competition between growing seeds on 1-year-old shoots and developing buds on current season growth. We studied the effects of chemical fruit thinning on alternate bearing and nut characteristics in a commercial orchard of ‘Owhadi’ pistachio cultivar during 2003–2004 in Rafsanjan, Iran. In both years, ethephon (100 and 200 mg l⁻¹), naphthaleneacetic acid (NAA) (125 and 250 mg l⁻¹) and urea (2.5 and 5%) were applied to the branch units of each individual ‘on’-year trees. The results showed that ethephon at both concentrations significantly increased fruit thinning and floral bud retention for the subsequent year. Other treatments also increased fruit thinning and floral bud retention but were inferior to ethephon.     

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.     

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.     

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.     

The fruit pitting disorder—A physiological anomaly in mango (Mangifera indica L.) due to deficiency of calcium and boron

A new disorder known as fruit pitting has been observed in some Indian mango orchards during the recent years. In this disorder, there is a development of some sunken pits on fruit peel, which distract consumers. Based on preliminary observations, it was observed that deficiency of nutrients could be the cause, and hence systematic studies were conducted in five indigenous cultivars such as ‘Alphonso’, ‘Amrapali’, ‘Dashehari’, ‘Mallika’ and ‘Neelum’, and five exotic mango cultivars such as ‘Edward’, ‘Irwin’, ‘Rosari’, ‘Sensation’ and ‘Tommy Atkins’ with the aim to observe the fruit pitting incidence and degree, and to investigate its probable causes. Our studies indicated that nearly 13% of the mango fruit was affected by fruit pitting with variable degree and magnitude. All indigenous cultivars had higher incidence of fruit pitting than exotic cultivars. ‘Dashehari’ had the maximum incidence of fruit pitting (30.3%), followed by ‘Amrapali’ (28.6%), and ‘Rosari’ the least (3.4%). Our studies indicated that the incidence of fruit pitting in mangoes was nearly 13% with a significant variability among the cultivars (Table 1). Although the concentrations of most of the major nutrients such as N, P, K, Mg, and minor nutrients such as Cu, Mn, Fe, Zn, did not differ significantly. However, the pitted fruit had lower Ca (1.53%) and B (22 mg kg⁻¹) concentrations than normal fruit (2.47% and 38 mg kg⁻¹, respectively), indicating that deficiency of Ca and B probably is the cause for fruit pitting in mangoes.     

Improving ‘Bing’ sweet cherry fruit quality with plant growth regulators

Final fruit diameter is the prime determinant of sweet cherry fruit value. Previous research has shown that mesocarp cell size accounts predominantly for variability in final fruit size, within a genotype. Our research program evaluated the potential to improve sweet cherry fruit size/weight with growth regulators to affect cell division and/or cell expansion stages. In the current study we screened 8 plant growth regulators (PGRs), including cytokinins, gibberellins, and auxins, and their combinations for their ability to increase ‘Bing’ fruit weight. Each PGR was mixed in lanolin paste and applied to fruit pedicels at 9 or 30 days after full bloom (DAFB), to coincide with estimated peak in cell division and cell expansion activity, respectively. Several cytokinins applied 30 DAFB improved fruit weight significantly (ca. +15%) with N-(2-Chloro-4-pyridyl)-N′-phenylurea (CPPU) and 6-(3-hydroxybenzylamino) purine (mt-Topolin) at 100 mg l⁻¹ being the most effective. Gibberellins, applied alone, improved fruit size and delayed fruit maturation and exocarp coloration. GA₃ at 200 mg l⁻¹ applied at 9 DAFB was the most effective and improved final fruit weight by 15%. Fifty-six percent of the fruit from this treatment were ≥9 g compared to 15% of similar weight fruit from untreated limbs. Both GA₃ and GA_4/7 treatments applied 9 DAFB increased fruit radial expansion. 4-Chlorophenoxyacetic acid, a synthetic auxin, also stimulated higher fruit growth rates at stage I and stage II, and fruit color development, but did not improve final fruit size.     

CPPU application on size and quality of hardy kiwifruit

For the purpose of determining the appropriate conditions of application to increase the size of a hardy kiwifruit, Actinidia arguta ‘Mitsuko’, N₁-(2-chloro-4-pyridyl)-N₃-phenylurea (CPPU) was applied at three different growth stages of the crop: at petal fall, 10 and 25 days after petal fall (DAPF), and three different concentrations: 1, 5 and 10 mg l⁻¹. A significant increase in fruit size was obtained by treatment at the concentrations of 5–10 mg l⁻¹ and at 10 DAPF. The fruit weight doubled. Although a significant reduction in the concentrations of total soluble solids (TSS), titratable acids (TA) and ascorbic acid (AsA) in the CPPU-treated fruits was recorded, the TSS/TA ratio and AsA content per fruit increased by the treatment. CPPU application at petal fall induced abnormally protruding fruit tip.     

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.     

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.     

Parthenocarpic fruit production in loquat (Eriobotrya japonica Lindl.) by using gibberellic acid

This study evaluates the effect of gibberellic acid (GA₃) in inducing parthenocarpy in ‘Algerie’ loquat, as well as the optimum treatment conditions and associated techniques, hand thinning and ringing, to produce seedless fruit with high enough quality for fresh consumption. GA₃ applied in the course of the phenological growth stages 504–508 of the BBCH-scale produced seedless fruits, with the magnitude of the response depending on the concentration applied and number of treatments. Percentage of panicles bearing seedless fruitlets significantly increased with increasing GA₃ concentrations up to 100 mg l⁻¹ and significantly and positively correlated with the number of treatments applied. Trees treated three times with 100 mg l⁻¹ developed more than 90% of panicles bearing almost 7 seedless fruits per panicle, which were smaller in size, drier and slightly acid but similar in TSS concentration and skin colour than seeded fruits from untreated trees. Fruit thinning to 3 fruits per panicle did not increase seedless fruit size, but ringing performed at the onset of cell enlargement stage, growth stage 702 of the BBCH-scale, significantly increased fruit size by 12–15%, depending on the year. Trees treated three times with 100 mg l⁻¹ of GA₃ and ringed produced 26 kg, on average, of seeded fruit of suitable commercial quality.     

High frequency embryogenic callus induction and plant regeneration from mature caryposis of big bluestem and little bluestem

Big bluestem (Andropogon gerardii Vitman) and little bluestem [Schizachyrium scoparium (Michaux) Nash.] are native to the North America and are important forage grasses and ornamental grasses. Both grasses are proposed as ideal biomass producers for cellulosic ethanol production. To apply genetic transformation, which is an important tool for incorporating desirable agronomic traits into plants to both species, however requires an efficient and reproducible regeneration protocol. We used mature caryopses from big and little bluestem as explants and tested the effect of various combinations of 2, 4-dichlorophenoxyacetic acid (2, 4-D) (1, 2, 3, 4 or 5 mg l⁻¹) and kinetin (KT) (0, 0.1 or 0.2 mg l⁻¹) on embryogenic callus induction with LS as the basal medium. The highest percentage of embryogenic calli induction occurred on medium containing 2, 4-D alone at 2 mg l⁻¹ for ‘Bison’ and on medium containing 4 mg l⁻¹ 2, 4-D alone for ‘Bonilla’ big bluestem. For little bluestem, the highest percentage of embryogenic callus induction occurred on medium containing 3 mg l⁻¹ 2, 4-D plus 0.1 mg l⁻¹ kinetin, suggesting that addition of KT is beneficial. Shoot regeneration took place on LS basal medium without any plant growth regulator for both species, although the addition of KT increased both regeneration frequency and the number of shoots produced per callus. Rooting of shoots reaching about 2 cm long occurred readily with or without α-naphthaleneacetic acid (NAA). Rooted plantlets were all successfully established in the soil.     

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.     

Effect of auxin treatments,cuttings’ collection date and initial characteristics on Paeonia ‘Yang Fei Chu Yu’ cutting propagation

Paeonia ‘Yang Fei Chu Yu’ is one of the most popular and commercially valuable cultivars of herbaceous peony. The study was performed to explore propagation techniques by cuttings for the nursery industry. Results showed that the stem cuttings pretreated with 2000 mg L⁻¹ indole-3-butyric acid (IBA) in quick-dip method got the best rooting traits (rooting percentage is 86.7%, root number is 23.1 and root length per rooted cutting is 6.4 cm). Therefore pre-treatment with 2000 mg L⁻¹ IBA is recommended.     

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%.     

Effect of genotype,explant size,position, and culture medium on shoot generation of Gerbera jamesonii by receptacle transverse thin cell layer culture

An unique procedure for the mass shoot propagation of Gerbera using receptacle transverse thin cell layer (tTCL) culture procedure was developed. Genotype, flower bud age, explant size, position of receptacle tTCLs and culture media were found to affect the success of culture. Ten interspecific crosses of Gerbera showed different shoot regeneration rates and callus induction via receptacle tTCL culture, all of which had shoot regeneration rates higher than 57%. Flower buds collected on the 10th day resulted in 91% shoot regeneration after 6 weeks of culture on basal MS medium [Murashige, T., Skoog, F., 1962. A revised medium for rapid growth and bioassay with tobacco tissue cultures. Physiol. Plant. 15, 475–497] supplemented with 0.02 mg l⁻¹ thidiazuron (TDZ), 0.8 mg l⁻¹ adenine and 10% (v/v) coconut water (CW). This was significantly higher than those from flower buds on the 7th and 14th days (22% and 54%), respectively. Shoot regeneration rate was the highest (94–100%) in the middle layers of the receptacle. For mass shoot propagation, shoot clusters were subcultured on half-strength MS medium supplemented with 0.5 mg l⁻¹ indole-3-butyric acid (IBA), 0.5 mg l⁻¹ 6-benzyladenine (BA) and 2.0 mg l⁻¹ kinetin after every 4 weeks. Plantlets formed when single shoots were cultured on half-strength MS medium containing 1 mg l⁻¹ IBA. All plantlets acclimatized well in the greenhouse.     

Direct plant regeneration from mature leaf explants of pistachio, Pistacia vera L.

A protocol was developed for direct shoot and plantlet regeneration from in vitro regenerated leaf explants of male Pistacia vera L. cv. ‘Atl?’. Leaves excised from axenic shoot cultures of pistachio were used to induce organogenesis on a Murashige and Skoog (MS) medium with Gamborg vitamins supplemented with combinations of different concentrations of BAP and IAA. The highest adventitious shoot regeneration in 35% of the explants, with the number of shoots ranging from 2 to 3 per explant, occurred in the explants cultured during the establishment phase in the medium with 1 mg l⁻¹ IAA and 2 mg l⁻¹ BAP. For shoot multiplication, the highest number of new microshoot/explants (5.76) was obtained in a culture medium supplemented with 1 mg l⁻¹ BAP, but it was not significantly different from the number obtained at 2 mg l⁻¹ BAP. A high rooting frequency (84%) for microshoots was recorded on a medium supplemented with 2 mg l⁻¹ IBA. In vitro rooted plantlets were transferred to pots filled with a mixture of soil, sand and peat (1:1:1). They were weaned in a growth room and finally moved to a greenhouse. This protocol could be utilized for in vitro clonal propagation of this economically important plant.