Rooting response of shoot cuttings from three peach growth habits

Current year shoot cuttings were collected in October and August from three growth habits of peach (Compact, Pillar, and Standard) and treated with one of four concentrations of indole butyric acid (0, 250, 1250, and 2500 mg L⁻¹ IBA). Rooting response was measured after 5 weeks in the greenhouse. Little or no rooting occurred with cuttings from any growth habit that was collected in October or in August when treated with 0 and 2500 mg L⁻¹ IBA. In August, the number of shoots that rooted was greater in cuttings from Pillar (79 and 45%) than Compact (13 and 3%) treated with 250 and 1250 mg L⁻¹ IBA, respectively. Cuttings from Standard trees had intermediate rooting of 56 and 6% at 250 and 1250 mg L⁻¹ IBA, respectively. Pillar trees consistently grew more roots with greater root length per cutting than the other growth habits. It is proposed that differences in rooting response among the growth habits may be associated with differences in endogenous auxin concentration that had been found in previous studies. Within peach and possibly other fruit trees, the capacity of shoot cuttings to develop adventitious roots can vary by cultivar and successful root induction with exogenous plant growth regulators may depend, in part, on endogenous hormone levels.     

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.     

Light penetration,growth, and fruit productivity in ‘Fuji’ apple trees trained to four growing systems

The study was conducted to document the range in soil nutrients, trunk and shoot growth, leaf characteristics and nutrient content, flower bud characteristics, yield, fruit quality, disease occurrence, light penetration, and shoot bark color in a 15-year-old ‘Fuji’ apple (Malus × domestica Borkh.) orchard on M.26 rootstocks trained to four growing systems. SS trees naturally growing had greater width and volume than the LT lightly heading, MLT heavily heading, and S trees thinning and bending. LT trees had more upward terminal shoots at old branches, large numbers of lateral shoots, and longer total shoot length. Leaf [N], [P], and [K] were the lowest for the MLT trees. SS and S trees had greater flower bud density and fruit yield, but LT trees had poor fruit color and less soluble solids and firmness. Percent light penetration into the canopy was the highest with the S system but not different from MLT or SS. The lowest was for the LT system, however, it was not statistically different from MLT or SS.     

Plant regeneration of an endangered medicinal plant Hydrastis canadensis L.

Goldenseal (Hydrastis canadensis L.) is an endangered medicinal plant used to treat sore eyes and mouths, cold and flu and also as a dye. The objective of this study was to develop an efficient in vitro propagation protocol for goldenseal. Significantly more shoots (26 shoots per leaf explants) were induced on a medium containing 2.5 μM thidiazuron (TDZ) and 5.0 μM 1-naphthaleneacetic acid (NAA) than any other treatment. Sub-culturing regenerated shoots on a medium with 5.0 μM 6-benzylaminopurine (BA) induced the maximum rate of shoot multiplication. Growth of the regenerated shoots in a temporary immersion bioreactor resulted in significant increases in biomass, shoot height and shoot multiplication. The regenerated shoots from the temporary immersion bioreactor formed roots when transferred onto a medium with 1.0–2.0 μM indole-3-butyric acid (IBA). Regenerated whole plantlets were acclimatized and maintained in standard greenhouse conditions for further growth. The regeneration protocol developed in this study provides a basis for germplasm conservation and for further investigation of this rare, medicinally important species.     

In vitro plantlets from alginate-encapsulated shoot tips of Solanum nigrum L.

Shoot tip explants obtained from in vitro proliferated shoots were encapsulated in 3% sodium alginate and 100 mM calcium chloride for the production of synthetic seed in Solanum nigrum L., a medicinally important plant. Morphogenic responses of encapsulated shoot tips to various sowing media (full or half-strength 0.8% agar-solidified or liquid MS medium or full-strength MS medium containing BAP) were evaluated in vitro. Of the six media evaluated, maximum conversion was obtained on 0.8% agar-solidified growth regulator free full-strength MS medium. The addition of MS nutrients in alginate matrix had a pronounced effect on the length of shoots that emerged from alginate beads. Encapsulated shoot tips also converted when directly sown in sterile soil moistened with liquid MS medium. Encapsulated shoot tips could be stored at low temperature (4 °C) up to 60 days. Plantlets regenerated from encapsulated shoot tips were acclimatized successfully.     

Effects of multiple applications of methyl jasmonate on fruit ripening,leaf gas exchange and vegetative growth in fruit trees

Summary

Multiple applications of methyl jasmonate (Me-J), at 10.mM, enhanced the rate of fruit ripening in peach ‘Redhaven’. The effect of Me-J on red colour development and fruit firmness was most pronounced in fruit treated six times with Me-J. Lower concentrations of Me-J had no effect of fruit colour. Me-J, at 10.mM, significantly reduced stomatal conductance, photosynthetic rate and transpiration rate of crabapple leaves, but had no effect on these parameters in peach leaves. The short-term inhibitory effects of Me-J on leaf gas exchange in crabapple persisted for up to 24 h. Fourteen applications of Me-J over a period of 28 days resulted in a 60% decrease in the concentration of chlorophyll in leaves. These Me-J applications also reduced the length of new branches, leaf number and leaf fresh weight by 65%, 31% and 47%, respectively. In peach, Me-J induced some leaf chlorosis and early leaf senescence within two weeks of the treatments which resulted in significant reductions in shoot length.     

Changes in morphological phenotypes and essential oil components in lavandin (Lavandula × intermedia Emeric ex Loisel.) transformed with wild-type strains of Agrobacterium rhizogenes

Hairy roots were induced from leaf-derived calli of lavandin (Lavandula × intermedia Emeric ex Loisel.) by infection with wild-type strains of Agrobacterium rhizogenes, A-5 (MAFF 02-10265) and A-13 (MAFF 02-10266). A-5-inoculated calli formed hairy roots more efficiently than A-13 ones. The transgenic shoots could be obtained from hairy root segments mediated by each Agrobacterium strain. However, different plant growth regulators were required for efficient adventitious shoot formation in each strain. In A-5, the most efficient adventitious shoot formation rate of 23.8% was observed in a medium with 4.4 × 10⁻⁶ M of 6-benzylaminopurine. On the other hand, a significantly higher rate of 13.2% was detected in a medium with 4.0 × 10⁻⁷ M of N-(2-chloro-4-pyridyl)-N′-phenylurea in A-13. Most of the regenerated plants showed dwarfism with closed internodes and extensive lateral branching, which were typical characteristics of ‘hairy root syndrome’. On the other hand, only nine of the 45 regenerated plants formed flower buds in early June, a delay of about one month compared with nontransgenic regenerated plants. The floral stalks and spikes of these plants were very short, resulting in a compacted form. Many regenerants showed a significantly lower productivity of essential oil than nontransgenic regenerants. Moreover, the relative percentage of the linalyl-cation-derived compounds, linalool and linalyl acetate, decreased in most of the regenerated plants. Compact plants with the ability of flower bud formation are assumed to be valuable not only for lavandin breeding, but also for clarifying the interaction between rol genes expression and essential oil production.     

In vitro shoot multiplication and plant regeneration in four Capsicum species using thidiazuron

A procedure of in vitro plant propagation using shoot meristem explants (∼0.5 cm) has been developed for Capsicum annuum cv CA960, C. baccatum, C. frutescens and C. praetermissum on Murashige and Skoog [Murashige, T., Skoog, F., 1962. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Plant Physiol. 15, 473–497] medium containing various cytokinins. Among various concentrations of cytokinins tested; adenine (Ad), N⁶-benzyladenine (BA), kinetin (Kn), zeatin and thidiazuron (TDZ) individually. TDZ regenerated maximum number (4.2–22.4) of shoots in all the Capsicum species tested. Multiple shoot elongation occurred upon transfer to BA (0.22 μM l⁻¹) + IAA (0.48 μM l⁻¹). Rooting of regenerated shoots was achieved on medium supplemented with 5.71 μM l⁻¹ indole-acetic acid (IAA). Rooting was observed in 72–94% of shoots obtained from TDZ-containing regeneration medium followed by elongation treatment in contrast to 8–22% of shoots without elongation treatment. Plantlets obtained from TDZ-containing media were normal diploid (2n = 24) and could readily be established in the soil under green house conditions with a survival frequency of 68–84%. Regenerated plants were developed into morphologically normal, fertile plants and able to set viable seeds.     

Comparison of shoot induction ability of different explants in herbaceous peony (Paeonia lactiflora Pall.)

Shoot induction ability of explants of herbaceous peony was investigated in semisolid MS medium containing BA, TDZ and GA₃. Callus was readily induced from stem without node and petiole explants within 2 days of culture but failed to generate shoots. Adventitious shoots were successfully produced from meristematic regions only: bud eyes on nodal stem sections, and junctions of petioles and petiolules. No shoots were induced from internode sections, petiole without junctions, or leaf sections. Nodal sections were the most efficient explants. There were up to 20 shoots in one explant generated within 20 days of culture. TDZ was more effective than BA to induce shoots. The 100% shoot induction rate was obtained in medium containing 0.1–3 mg L⁻¹ of TDZ. However, higher concentrations of TDZ inhibited shoot elongation and only large leaf clusters were produced. Combinations of BA and TDZ failed to increase shoot induction rates but caused shoots shorter. The 2–60-min pretreatment of explants with 20 mg L⁻¹ TDZ solution was very effective to induce adventitious shoots directly, but both shoot number and shoot length tended to decrease as treatment time increased. GA₃ was beneficial for shoot and stem elongation.     

Mobilization of nitrogen in the olive bearing shoots after foliar application of urea

Mature ‘on’ and ‘off’ ‘Manzanillo’ olives trees with three healthy branches of 10–12 cm in diameter were selected to determine changes in nitrogen levels in the bearing shoots after foliar application of urea. Each selected branch of each tree received one of the following treatments: (i) control without urea application; (ii) foliar application of urea to all the current-season leaves; and (iii) foliar application of urea to all the one-year-old leaves. Urea was applied in May, four days after full bloom in the ‘on’ year trees. Each treated leaf was immersed in a test tube containing a 4% urea solution and 0.1% Tween 20 for 5 s. Bearing shoots, composed of both one-year stems and leaves and current-season stems and leaves, were collected at intervals from the beginning of the experiment until 64 days after urea application. Nitrogen was determined in stems and leaves from each part of the bearing shoot, and in fruits during the ‘on’ year. Nitrogen uptake from the leaves was rapidly mobilized from the older to the current-season leaves of the bearing shoot, and thereafter to other storage organs of the tree or to the fruit, which is the largest nitrogen sink in the bearing shoot. No translocation of nitrogen from the current-season leaves to older leaves was observed. The rapid translocation of nitrogen from the younger leaves to other storage organs of the tree could explain the insensitivity of leaf analysis to detect excess nitrogen, since mature leaves from current-season shoots must be sampled to determine the nutritional status of the tree. The failure of leaf analysis to detect excess nitrogen may be a cause of nitrogen over-fertilization in olive orchards.     

Cryopreservation of Malus cultivars: Comparison of two droplet protocols

Two droplet procedures, droplet-vitrification (PVS2) and droplet (DMSO) were applied for cryopreservation of in vitro cultured apple (Malus domestica Borkh., cvs. Florina, Idared, Colmar and Rebra) plants. The highest post-thaw regrowth rates (70% for cv. Florina, 66% for cv. Idared, 63% for cv. Colmar and 60% for cv. Rebra) were achieved after using the droplet-vitrification (PVS2) protocol. The excised shoot tips (2–3 mm in length) were precultured in 0.5 M sucrose enriched media for 24 h. Subsequently they were transferred in PVS2 vitrification solution for 30 or 40 min (depending on cultivar) at 24 °C and then immersed in liquid nitrogen. Rewarming was performed in liquid MS medium at 24 °C. Plants regenerated from cryopreserved shoot tips did not display any sign of morphological alteration or abnormalities in growth in comparison with control plants.     

High frequency shoot regeneration from cotyledon explants of cucumber via organogenesis

Organogenic callus induction and high frequency shoot regeneration were achieved from cotyledon explants of cucumber. About 86.2% of cotyledon explants derived from 5-day-old in vitro raised seedlings produced green, compact nodular organogenic callus in MS medium containing NAA (2.69 μM) and BA (4.44 μM) after two successive transfers at 20 days interval. Adventitious shoots were produced from the organogenic callus when it was transferred to MS medium supplemented with NAA (1.34 μM), BA (8.88 μM), zeatin (0.91 μM) and l-glutamine (136.85 μM) with shoot induction frequency of 75.6%. Shoot proliferation occurred when callus with emerging shoots was transferred in the same medium at an interval of 20 days. Shoots (1.0 cm length) were excised from callus and were elongated in MS medium fortified with GA₃ (1.44 μM) and BA (4.44 μM). The elongated shoots were rooted in MS medium supplemented with IBA (3.42 μM) and BA (4.44 μM). Rooted plants were acclimatized in green-house and subsequently established in soil with a survival rate of 80%. This protocol yielded an average of 35 shoots per cotyledon explant in a culture duration of 120–140 days.     

Gibberellic acid and flower bud development in loquat (Eriobotrya japonica Lindl.)

The application of gibberellic acid (GA₃) to the whole loquat tree from mid-May to early June and from early August to the onset of flowering, significantly reduced the number of premature flowering shoots per current shoot and per m³ of canopy, and so reduced the total number of panicles per m³ of canopy. The number of vegetative shoots per m³ of canopy was also significantly reduced by applying GA₃. The response depended on the concentration applied and produced optimal results at 50 mg l⁻¹. Differences in the number of flowers per panicle and leaves per shoot were not significantly modified by the treatment. Nevertheless, GA₃ applied directly to the developing apex near to flower differentiation reduced the number of flowers per panicle by 25–35% and without modifying the morphological characteristics of the panicle. Results suggest that less sprouting of lateral buds was largely responsible for the reduction in flowering intensity caused by GA₃. Best treatments reduced thinning costs (60%, approximately) of premature flowering shoots, slightly increased fruit diameter and significantly improved fruit colour and juice quality, thus advancing harvest date.     

Partitioning of C-photosynthates from different current shoots neighboring with fruiting spur in later-maturing Japanese pear during the period of rapid fruit growth

To elucidate the fate of photosynthates from different current shoots and their influence on fruit growth and bud differentiation in neighboring spur complex during the period of rapid fruit growth in two late-maturing Japanese pear cultivars: ‘Atago’ and ‘Shinkou’ with contrasting fruit size, ¹³C labeling of single shoot was done to investigate of C-relations in fruit branches of eight shoot-combinations. The results showed that all of the current shoots investigated (bourse shoots of nonfruiting spur, bourse shoots of fruiting spur, extension shoot, nonfruiting spur, vegetative shoot, and water sprout) could export photosynthates to the neighboring fruit and buds. Water sprouts together with vegetative shoots, bourse shoots, and extension shoots are important source for fruit growth after shoot growth termination during the period of rapid fruit growth in production of late-maturing pears. The carbon transfer rate from the neighboring to the fruit bearing spur is depent (i) on the types of shoot which acts as C source, (ii) on the position of the fruiting spur and (iii) on the source-sink distance. Furthermore, the cultivar difference in carbon partitioning from different current shoot-combinations confirmed that the movement of photosynthates into the fruit was determined by sink strength of the fruit, and ‘Atago’ exhibited a greater relative sink strength of fruit than ‘Shinkou’. In addition, vegetative shoots are very important C sources for fruit growth in ‘Atago’ and the growth pattern of bourse shoot seriously affects C allocated to fruit in ‘Shinkou’.     

Micrografting of almond (Prunus dulcis Mill.) cultivars “Ferragnes” and “Ferraduel”

The success of various in vitro micrografting techniques, establishment of the rootstock, size of the microscion, and the effects of culture medium on the grafted seedling development for almond cultivars “Ferragnes” and “Ferraduel” were studied. In vitro germinated wild almond seedlings developed from seeds were used as rootstocks. Shoot culture initiation was successfully achieved from the above almond cultivars by culturing mature shoot tips from forced nodal buds, about 3–5 mm, on 0.7 mg/L BA and 0.01 mg/L NAA containing a MS medium. The regenerated adventitious shoots from in vitro cultures were maintained and proliferated by sub-culturing on a fresh medium every three to 4 weeks. Regenerated shoot tips, which were micrografted onto in vitro seedlings, resulted in the restoration of shoot proliferation. The results indicated that the most successful method for the grafting of tested almond cultivars was slit micrografting. High levels of micrograft take were achieved with all ranges of scions (4–15 mm) obtained from the regenerated shoot tips. Slow growth and lack of axillary shoot development on the micrografts were noticeable when the micrografts were cultured on hormone-free germination medium. In vitro micrografted plantlets were successfully acclimatized and no problems were encountered with the establishment of micrografted plants in vivo. The developed technique has demonstrated a high potential for application in the micropropagation of almond cvs. “Ferragnes” and “Ferraduel” and thereby, represents a feasible method for the renewal of almond orchards in Turkey and elsewhere in the world.     

In vitro germination and plantlet establishment of Labisia pumila (Bl.) F. Vill.

In vitro seeds germination and plantlet establishment of Labisia pumila were studied in this report. The seeds obtained from the mature fruits of L. pumila were sterilized and cultured on Murashige and Skoog (MS) solid media supplemented with 1–3 μM of 6-benzylaminopurine (BAP) and 3% (w/v) sucrose. The presence of BAP in the medium significantly affects seeds germination. High percentage of seeds germination (up to 90%) was successfully achieved after 2 weeks of culture on medium supplemented with 2 μM BAP. Up to 70% of explants produced shoots through direct regeneration from newly emerged epicotyls after 5 weeks of culture. The average of 8.1 ± 1.0 shoots per explant obtained on media treated with 2 μM BAP. Seedlings were further transferred to growth media fortified with different types of cytokinin. Result observed after 12 weeks showed that medium supplemented with 1 μM zeatin (ZEA) promote the highest growth with an average of 2.9 ± 1.0 cm shoot length and 7.7 ± 3.2 leaves per explant after 12 weeks. In addition, medium added with 2 μM BAP and supplemented with 3–4% (w/v) of sucrose promote the best growth i.e., 3.0 ± 0.6 shoots per explant, 2.27 ± 0.2 cm length and 4.3 ± 0.5 leaves per explant.     

Horticultural factors regulating effectiveness of GA3 inhibiting flowering in peaches and nectarines (Prunus persica L. Batsch)

The application of gibberellic acid during the flower bud induction period significantly reduced flowering in peaches and nectarines. The magnitude of the response significantly depended on the total amount of active material applied per tree. Results show, for cultivars tested, a higher sensitivity of peach to GA₃ in comparison with nectarine. Concentrations of 0.5 or 1.0 g tree⁻¹ of gibberellic acid reduce flowering by about 50% in both, peaches and nectarines, respectively, and it gives rise to a reduction of costs of hand thinning by 50%, approximately, without affecting the yield. Fruit colour advanced, total soluble solids concentration increased and fruit firmness increased as a consequence of treatments. The effect was higher in the basal part of the shoots and reduced from the base to the apical part.     

The Effect of Partial Defoliation at Different Times in the Season on Fruit Drop and Shoot Growth in Lord Lambourne Apple Trees

Six-branched trees were deblossomed on alternate branches and various defoliation treatments given. For the period of 7 to 30 days from the time of treatment, defoliations increased the compound-interest rates of fruit drop. After this period these rates of drop became equal to those of untreated trees and the relative differences in crop numbers established by this time persisted until harvest though the actual numbers still fell. The earlier in the season that the defoliations were applied the greater were the relative differences. The removal of spur leaves from branches bearing fruit caused greater drop than the removal of spur leaves from neighbouring deblossomed branches. The removal of leaves from extension and bourse shoots of fruiting or deblossomed branches had no significant effect upon fruit drop or shoot growth.

The removal of all flowers from alternate branches at mouse-ear caused more shoots to grow from these branches than from branches allowed to fruit. Removal, at pink bud, of spur leaves from either fruiting or non-fruiting branches increased the number of shoots on both branches, but removal deferred till later in the season had no effect.     

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.