Growth and Morphogenesis in Tissue Cultures of Anagallis arvensis

Morphogenesis has been induced in excised organs and callus tissue cultures obtained from various parts of the seedling and mature plants of pimpernel (Anagallis arvensis). Vigorously growing cell cultures capable of being periodically subcultured have been established in liquid as well as on the agar-solidified Murashige and Skoog's medium supplemented with 2,4-D (0.1 mg/1) + kinetin (0.1 mg/l) + coconut milk (10%). The callus tissue obtained from excised hypocotyl segments is white, soft, friable and fast growing, and has been subcultured over a period of two years without showing any sign of decline in growth. The optimum conditions for growth are at pH 5.9, temperature 27°C, and with 4% sucrose as the carbon source. Under appropriate nutritional supply these cultures can be manipulated to induce rhizogenesis in the suspension cultures, and buds and “embryo-like” structures on agar-solidified media. The excised leaves, hypocotyl and stem segments regenerate buds. Of the cytokinins used, 6-(y,y-dimethylallylamino)-purine proved to be the best for the number of cultures producing buds, as well as for the number of buds per culture. Anatomical studies revealed that buds arise from the epidermal and subepidermal layers of leaves and hypocotyl; these buds form shoots which eventually develop into plantlets.     

Effect of Assimilate Supply on Development and Growth Potential of Axillary Buds in Roses

The effect of assimilate supply on axillary bud developmentand subsequent shoot growth was investigated in roses. Differencesin assimilate supply were imposed by differential defoliation.Fresh and dry mass of axillary buds increased with increasedassimilate supply. The growth potential of buds was studiedeither by pruning the parent shoot above the bud, by graftingthe bud or by culturing the bud in vitro. Time until bud breakwas not clearly affected by assimilate supply during bud development,Increase in assimilate supply slightly increased the numberof leaves and leaf primordia in the bud; the number of leavespreceding the flower on the shoot grown from the axillary budsubstantially increased. No difference was found in the numberof leaves preceding the flower on shoots grown from buds attachedto the parent shoot and those from buds grafted on a cutting,indicating that at the moment of release from inhibition thebud meristem became determined to produce a specific numberof leaves and to develop into a flower. Assimilate supply duringaxillary bud development increased the number of pith cells,but the final size of the pith in the subsequent shoot was largelydetermined by cell enlargement, which was dependent on assimilatesupply during shoot growth. Shoot growth after release frominhibition was affected by assimilate supply during axillarybud development only when buds sprouted attached to the parentshoot, indicating that shoot growth is, to a major extent, dependenton the assimilate supply available while growth is taking place.Copyright1994, 1999 Academic Press Assimilate supply, axillary bud, cell number, cell size, defoliation, development, growth potential, meristem programming, pith, Rosa hybrida, rose, shoot growth     

Changes in the Endogenous Cytokinins of Bark and Buds ofSalix babylonica as a Result of Stem Girdling

Girdling of 1-year-old Salix babyionica L. plants resulted in an early accumulation of compounds which co-chromatographed with cytokinin glucosides in both the bark and buds below the girdle. In the bark the cytokinin glucosides were present in high levels in both girdled and non-girdled plants. In the buds of non-girdled plants. however, glucoside concentration was initially low but then increased rapidly after ringing and reached a maximum level prior to any visible signs of bud swell. With the onset of lateral shoot growth the glucoside cytokinins decreased while the cytokinins that co-chromatographed with zeatin and its derivatives increased. As the cytokinin glucosides are generally considered to be storage forms, their accumulation in the bark and buds below the girdle apparently does not reflect synthesis but rather transport towards a more competitive sink. In the case of Salix plants the lateral buds would appear to have the ability to hydrolyze these glucosylated zeatin derivatives and then to utilize them for bud development. It is suggested that in the presence of a functional root system lateral buds do not synthesize cytokinins de novo, but that they do have the metabolic capacity to convert cytokinins transported to them.     

Micropropagation of adult Swainsona formosa (Leguminosae: Papilionoideae: Galegeae)

Summary Explants of axillary buds excised from mature adult stems of Swainsona formosa (G. Don) J. Thompson (syn. Clianthus formosus) were cultured on Murashige and Skoog medium supplemented with a range of auxins, cytokinins, and sucrose concentrations. Auxins did not increase shoot or bud numbers above controls, and 2,4-dichlorophenoxyacetic acid was the only auxin to significantly increase callus production. Benzyladenine or thidiazuron incorporated into the medium at 0.1 μM stimulated shoot and bud production, and shoot growth occurred following removal of cytokinins from the medium after 4 wk. Shoot number increased linearly with sucrose concentration up to 40 g l⁻¹, but shoot height and the number of cytokinin-induced buds were optimal at sucrose levels of 20–30 g l⁻¹. Roots were initiated in vitro following treatment of cuttings with 0.1% indole-3-butyric acid and 0.1% α-naphthaleneactic acid. Plantlets were successfully established in soil but were plagiotropic and exhibited distichous phyllotaxy.     

Effect of auxins and cytokinins on plant regeneration from hypocotyls and cotyledons in niger (Guizotia abyssinica Cass.)

Tissue cultures were established from hypocotyl and cotyledonary leaf segments ofGuizotia abyssinica Cass. on MS medium supplemented with various concentrations of auxins (IAA, NAA, IBA or 2,4-D) and cytokinins (KN or BA). Expiants cultured on media with cytokinins or in combination with auxins produced shoot buds. Maximum number of shoot buds (20–25 per culture) were differentiated from cotyledonary leaf segments on medium with 2 mg 1^-1 each of KN and IBA. Rooting of regenerated shoot buds was acheived on medium with NAA. The obtained plantlets were successfully transferred to soil.     

DIFFERENTIATION OF LATERAL SHOOTS AS THORNS IN ULEX EUROPAEUS

Ulex europaeus is a much-branched shrub with small, narrow, spine-tipped leaves and axillary thorn shoots. The origin and development of axillary shoots was studied as a basis for understanding the changes that occur in the axillary shoot apex as it differentiates into a thorn. Axillary bud primordia are derived from detached portions of the apical meristem of the primary shoot. Bud primordia in the axils of juvenile leaves on seedlings develop as leafy shoots while those in the axils of adult leaves become thorns. A variable degree of vegetative development prior to thorn differentiation is exhibited among these secondary thorn shoots even on the same axis. Commonly the meristems of secondary axillary shoots initiate 3–9 bracteal leaves with tertiary axillary buds before differentiating as thorns. In other cases the meristems develop a greater number of leaves and tertiary buds as thorn differentiation is delayed. The initial stages in the differentiation of secondary shoot meristems as thorns are detected between plastochrons 10–20, depending on vigor of the parent shoot. A study of successive lateral buds on a shoot shows an abrupt conversion from vegetative development to thorn differentiation. The conversion involves the termination of meristematic activity of the apex and cessation of leaf initiation. Within the apex a vertical elongation of cells of the rib meristem initials and their immediate derivatives commences the attenuation of the apex which results in the pointed thorn. All cells of the apex elongate parallel to the axis and proceed to sclerify basipetally. Back of the apex some cortical cells in which cell division has persisted longer differentiate as chlorenchyma. Although no new leaves are initiated during the extension of the apex, provascular strands are present in the thorn tip. Fibrovascular bundles and bundles of cortical fibers not associated with vascular tissue differentiate in the thorn tip and are correlated in position with successive incipient leaves in the expected phyllotactic sequence, the more developed bundles being related to the first incipient leaves. Some secondary shoots displayed variable atypical patterns of meristem differentiation such as abrupt conversion of the apex resulting in sclerification with limited cell elongation and small, inhibited leaves. These observations raise questions concerning the nature of thorn induction and the commitment of meristems to thorns.     

<Emphasis Type="Italic">In vitro</Emphasis> adventitious shoot bud differentiation and plantlet regeneration in <Emphasis Type="Italic">Feronia limonia</Emphasis> L. (Swingle)

Summary In vitro adventitious shoot bud regeneration systems are considered most suitable for Agrobacterium-and biolisticsmediated genetic transformation to obtain transgenic plants. In the present investigation, multiple adventitious shoot buds could be induced directly from Feronia limonia hypocotyl explants inoculated on Murashige and Skoog (MS) medium containing different growth regulators. During the initial phase, the hypocotyl segments nearer to the cotyledons responded quickly compared to those closer to the root. The response, however, was comparable in both the segments in subsequent subculture. Of the various cytokinins, 2.22 μM 6-benzylaminopurine (BA) proved to be more effective compared to kinetin (Kn). The two-way interaction of BA and Kn significantly influenced shoot regeneration and contributed the most among the interactions studied. The best response, however, was obtained when 2.22 μM BA and 2.32 μM Kn were combined. Although the effect of auxins like α-naphthaleneactic acid (NAA) combined with cytokinins evoked a significant responsein terns of number of shoot buds, this response did not supersede the effect of combined cytokinins. Vone of the polyamines tested induced shoot buds on hypocotyl segments. Adventitious shoots were multiplied on MS medium containing 2.22 μM BA, 6.96 μM Kn, and 0.05 μM NAA. More than 60% of the shoots produced roots when cultured on medium containing one-quarter strength MS salts, 10% suerose, 0.6% agar, and 7.36μM indole-3-butyric acid. The adventious origin of shoot buds showing continuous vascular connections was confirmed through histological investigations.     

Role of polarity in de novo shoot bud initiation from stem disc explants of <Emphasis Type="Italic">Curculigo orchioides</Emphasis> Gaertn. and its encapsulation and storability

The occurrence of strong polarity towards shoot bud induction and the effect of cytokinin(s) on each segment of stem axis, encapsulation and storability of de novo Shoot buds of Curculigo orchioides Gaertn. (Hypoxidaceae) have been documented in the present communication. Maximum number of shoot buds arising de novo from the stem discs (cross section) explanted from proximal end on MS medium fortified with BAP and KIN 1 mg/L each. Stem discs from distal end were less efficient in shoot bud induction. A combination of two cytokinins (BAP and KIN) as a synergistic effect on shoot buds induction from each segment of stem axis. Stem discs in inverted position produced shoot buds from the lower surface, showing strong polarity within the explant. Further, storability and shoot development of sodium alginate encapsulated shoot buds of Curculigo orchioides were tested on half-strength Murashige and Skoog (MS) basal medium fortified with coconut water (10% v/v). The frequency of regeneration from encapsulated shoot buds was affected significantly by concentration of sodium alginate and the duration of exposure to calcium chloride. Shoot buds encapsulated with 2.5% sodium alginate dissolved in MS basal salts solution recorded significantly higher shoot development than other treatments. A relatively short (5 min) incubation with calcium chloride solution provided uniform encapsulation of shoot buds that gave the highest percentage (68%) of shoot development. Encapsulated shoot buds could be stored at 4°C for 50 days without reduction in viability as oppose to non-encapsulated shoot buds, which showed 9.5% viability after 20 days at 4°C. Encapsulated shoot bud developed into normal shoots. Based on the present observations an improved protocol may be developed for the rapid multiplication and conservation of the endangered species—C. orchioides.     

Comparative studies of cytokinins on in vitro propagation of Bacopa monniera

A mass in vitro propagation system for Bacopa monniera (L.) Wettst. (Scrophulariaceae), a medicinally important plant, has been developed. A range of cytokinins have been investigated for multiple shoot induction with node, internode and leaf explants. Of the four cytokinins (6-benzyladenine, thidiazuron, kinetin and 2-isopentenyladenine) tested thidiazuron (6.8 μM) and 6-benzyladenine (8.9 μM) proved superior to other treatments. Optimum adventitious shoot buds induction occurred at 6.8 μM thidiazuron where an average of 93 shoot buds were produced in leaf explants after 7 weeks of incubation. However, subculture of leaf explants on medium containing 2.2 μM benzyladenine yielded a higher number (129.1) of adventitious shoot buds by the end of third subculture. The percentage shoot multiplication (100%) as well as the number of shoots per explant remained the high during the first 3 subculture cycles, facilitating their simultaneous harvest for rooting. In vitro derived shoots were elongated on growth regulator-free MS medium and exhibited better rooting response on medium containing 4.9 μM IBA. After a hardening phase of 3 weeks, there was an almost 100% transplantation success in the field. This revised version was published online in June 2006 with corrections to the Cover Date.     

Further studies on branch buds in mosses; “Pseudoparaphyllia” and “scaly leaves”

The development of so-called foliose pseudoparaphyllia in the species of theClimacium-type branch development was studied with a paraffin sectioning method and SEM. Scaly leaves (or scale-like leaves) and “foliose” pseudoparaphyllia proved to originate as leaves by segmentation of an apical cell of a branch initial in the very first stage of development into a branch bud. Branch buds of two species among the six species examined develop linear-lanceolate appendages which serve to protect the buds as well as scaly leaves. These appendages originate in the peripheral part of the epidermal layer of buds, and therefore they can be homologous to trichomes previously reported for species with branch primordia. Emendation of two terms are proposed; every organ originating from buds as leaves that are effective for bud protection should be called scaly leaves, while those which homologous to trichomes should be called pseudoparaphyllia. Both terms are used here in a narrower sense than before: There might befilamentous orfoliose scaly leaves, andfilamentous orfoliose (linear-to broad-lanceolate) pseudoparaphyllia. A scheme is given to show the pattern of branch development and the manner in which branch buds (or primordia) are protected.     

In vitro studies on the anatomy and morphology of bud regeneration in melon cotyledons

Summary The anatomy and morphology of bud regeneration were investigated in melon (Cucumis melo L.) cv. Galia, which regenerates in vitro only by direct organogenesis from the cotyledon explant. Explants were cut from the cotyledon proximal to the apex from 3-d-old in vitro seedlings. After 3 d on Murashige and Skoog medium with N⁶-benzyladenine, cell division can be observed in the epidermal layer on the adaxial side in the center of the explant, near the most proximal (wounded) cut edge. Over the next week, the area of the meristem increases laterally. Additional cell layers are added to the meristematic area by cell division in the epidermis. In places the epidermis remains active in cell division. Alongside those active areas there are zones where the epidermis has become inactive, although the subepidermal layers continue to divide. In transverse section, the explant now has small protuberances on the adaxial surface. After 10 d on cytokinin-containing medium, the first signs of development are visible on the adaxial surface adjacent to the proximal cut edge. The protuberances observed after 10 d are neither primordia nor buds, although some meristematic bulges are observed. The first regenerated shoot buds are observed histologically after 15 d, by which time the surface has many protuberances and some small leaves. The first shoot is found by histology after 22 d. By this time the surface is covered with protrusions and leaves, mostly without accompanying buds. The leaves may be produced from the protrusions initially visible after 10 d.     

Plantlet regeneration from fascicular buds of seedling shoot apices of <Emphasis Type="Italic">Pinus roxburghii</Emphasis> Sarg

Shoot apices of Pinus roxburghii Sarg were cultured on Murashige and Skoog’s medium (MS) supplemented with cytokinins [6-benzyladenine (BA), kinetin and N-benzyl-9-(2-tetrahydropyranyl) adenine (BPA)] alone and in combination with auxin, α-napthaleneacetic acid (NAA). Of the three cytokinins tested at varying concentrations, medium supplemented with 10 μM BA was found optimal in respect of explant responsiveness (97.22 %) and average number of buds induced per explant (7.42). The concentration of cytokinins in the induction medium had a profound effect on rate of elongation of induced buds on MS basal medium containing 0.5 % activated charcoal. Further, shoots induced on lower concentrations of BA increased up to 2.4 times in length in 4 weeks. Decapitation of the explant enhanced the rate of axillary bud elongation. Proliferating shoot cultures were established by sub-culturing the axillary shoots on MS supplemented with 10 μM BA. Shoots 2–3 cm in length were suitable for culturing as more buds were induced on them compared to longer or shorter shoots. Root primordia were induced on 70.83 % shoots when transferred to 1/2 MS medium supplemented with 5.0 μM NAA. Elongation of root primordia (60 %) was achieved in liquid 1/2 MS basal medium. The plantlets were successfully transferred to soil after hardening; the time period from initiation of shoot buds to transplantation being 20–22 weeks.     

Identification and quantification of the major endogenous cytokinins in pistachio seedlings

The major endogenous cytokinins, Z, ZR, DHZ, DHZR, iP and iPR in pistachio seedlings (Pistacia vera L. cv. Ohadi) were purified by HPLC and their identities confirmed using GC-MS. The aerial parts of two-year old pistachio seedlings including mature leaves, young leaves, lateral buds, debarked stems and bark were subjected to analysis. All of the above mentioned cytokinins were identified in the aerial parts except DHZ which was only present in mature leaves. Z-type cytokinins contributed almost 43% of the total cytokinins. ZR and DHZR were identified as the major ribosides and iP as the main base. The greatest concentration of ZR was detected in the bark, amounting to about 48%. DHZR and ZR constituted the major portion of the total cytokinins detected in both young and mature leaves while Z was detected as a minor cytokinin in leaves. The sharp increase of iP concentration during leaf maturation indicates that mature leaves are probably capable of de novo biosynthesis of cytokinins. The absence of DHZ (except in mature leaves) and the presence of considerable concentrations of DHZR in pistachio stems suggest that these tissues are able to metabolize DHZ to DHZR. The large amount of ZR in pistachio leaves suggests that root-derived ZR is transported into the leaves after loading into the xylem. The presence of high amounts of iP in pistachio lateral buds indicates that iP has been accumulated in these parts. The occurrence of a totally different cytokinin distribution pattern in buds, as compared with the other aerial parts, possibly results from their different metabolism.     

Root-synthesized cytokinin in Arabidopsis is distributed in the shoot by the transpiration stream

To clarify how root-synthesized cytokinins (CKs) are transported to young shoot organs, CK distribution patterns were analysed in free-CK-responsive ARR5::GUS transformants of Arabidopsis thaliana (L.) Heynh. together with free plus bound CKs using specific CK monoclonal antibodies. Plants were subjected to two different growth conditions, completely protected from any air movement, or exposed to gentle wind 3 h before harvesting. In wind-protected plants the strongest ARR5::GUS expression was found in the root cap statocytes, spreading upwards in the vascular cylinder. This pattern in roots was congruent with that found by CK immunolocalization. Shoots of wind-protected plants displayed either no or only low ARR5::GUS expression in the stem vascular bundles, nodal ramifications, and the bases of flower buds; shoot vascular bundles showed patterns of acropetally decreasing staining and the apical parts of buds and leaves were free from ARR5::GUS expression. In wind-exposed plants ARR5::GUS expression was considerably increased in shoots, also in basal-to-apical decreasing gradients. Immunolabelled shoots showed differential staining, with the strongest label in the vascular bundles of stems, leaves, and buds. The fact of the apparent absence of free CK in the buds of wind-protected plants and the typical upward decreasing gradients of free and conjugated CKs suggest that the bulk of the CK is synthesized in the root cap, exported through the xylem and accumulates at sites of highest transpiration where cuticles do not yet exist or do not protect against water loss.     

Correlation of Endogenous Gibberellic Acid with Initiation of Mango Shoot Growth

Stems of mango (Mangifera indica L.) rest in a nongrowing, dormant state for much of the year. Ephemeral flushes of vegetative or reproductive shoot growth are periodically evoked in apical or lateral buds of these resting stems. The initiation of shoot growth is postulated to be primarily regulated by a critical ratio of root-produced cytokinins, which accumulate in buds and by leaf-produced auxin, which decreases in synthesis and transport over time. Exogenously applied gibberellic acid (GA3) delays initiation of bud break but does not determine whether the resulting flush of growth is vegetative or reproductive. We tested the hypothesis that endogenous GA3, which influences release of these resting buds, may decrease in stem tips or leaves with increasing age of mango stems. GA3 and several other GAs in stem tip buds and leaves were identified and quantified in stems of different ages. The major endogenous GAs found in apical buds and leaves of vegetative mango stems were early 13-hydroxylation pathway gibberellins: GA1, epi-GA1, GA3, GA19, GA20, and GA29, as identified by gas chromatography-mass spectrometry (GC-MS). A novel but unidentified GA-like compound was also present. The most abundant GAs in apical stem buds were GA3 and GA19. Contrary to the hypothesis, the concentration of GA3 increased within buds with increasing age of the stems. The concentrations of other GAs in buds were variable. The concentration of GA3 did not change significantly with age in leaves, whereas that of most of the other GAs declined. GA1 levels were greatest in leaves of elongating shoots. These results are consistent with the concept that rapid shoot growth is associated with synthesis of GAs leading to GA1. The role of GA3 in delaying bud break in mango is not known, but it is proposed that it may enhance or maintain the synthesis or activity of endogenous auxin. It, thereby, maintains a high auxin/cytokinin ratio similar to responses to GA3 that maintain apical dominance in other plant species.     

Correlation of Endogenous Gibberellic Acid with Initiation of Mango Shoot Growth

Stems of mango (Mangifera indica L.) rest in a nongrowing, dormant state for much of the year. Ephemeral flushes of vegetative or reproductive shoot growth are periodically evoked in apical or lateral buds of these resting stems. The initiation of shoot growth is postulated to be primarily regulated by a critical ratio of root-produced cytokinins, which accumulate in buds and by leaf-produced auxin, which decreases in synthesis and transport over time. Exogenously applied gibberellic acid (GA3) delays initiation of bud break but does not determine whether the resulting flush of growth is vegetative or reproductive. We tested the hypothesis that endogenous GA3, which influences release of these resting buds, may decrease in stem tips or leaves with increasing age of mango stems. GA3 and several other GAs in stem tip buds and leaves were identified and quantified in stems of different ages. The major endogenous GAs found in apical buds and leaves of vegetative mango stems were early 13-hydroxylation pathway gibberellins: GA1, epi-GA1, GA3, GA19, GA20, and GA29, as identified by gas chromatography-mass spectrometry (GC-MS). A novel but unidentified GA-like compound was also present. The most abundant GAs in apical stem buds were GA3 and GA19. Contrary to the hypothesis, the concentration of GA3 increased within buds with increasing age of the stems. The concentrations of other GAs in buds were variable. The concentration of GA3 did not change significantly with age in leaves, whereas that of most of the other GAs declined. GA1 levels were greatest in leaves of elongating shoots. These results are consistent with the concept that rapid shoot growth is associated with synthesis of GAs leading to GA1. The role of GA3 in delaying bud break in mango is not known, but it is proposed that it may enhance or maintain the synthesis or activity of endogenous auxin. It, thereby, maintains a high auxin/cytokinin ratio similar to responses to GA3 that maintain apical dominance in other plant species.     

Development and Growth Potential of Axillary Buds in Roses as Affected by Bud Age

The effect of axillary bud age on the development and potentialfor growth of the bud into a shoot was studied in roses. Ageof the buds occupying a similar position on the plant variedfrom 'subtending leaf just unfolded' up to 1 year later. Withincreasing age of the axillary bud its dry mass, dry-matterpercentage and number of leaves, including leaf primordia, increased.The apical meristem of the axillary bud remained vegetativeas long as subjected to apical dominance, even for 1 year. The potential for growth of buds was studied either by pruningthe parent shoot above the bud, by grafting the bud or by culturingthe bud in vitro. When the correlative inhibition (i.e. dominationof the apical region over the axillary buds) was released, additionalleaves and eventually a flower formed. The number of additionalleaves decreased with increasing bud age and became more orless constant for axillary buds of shoots beyond the harvestablestage, while the total number of leaves preceding the flowerincreased. An increase in bud age was reflected in a greaternumber of scales, including transitional leaves, and in a greaternumber of non-elongated internodes of the subsequent shoot.Time until bud break slightly decreased with increasing budage; it was long, relatively, for 1 year old buds, when theysprouted attached to the parent shoot. Shoot length, mass andleaf area were not clearly affected by the age of the bud thatdeveloped into the shoot. With increasing bud age the numberof pith cells in the subsequent shoot increased, indicatinga greater potential diameter of the shoot. However, final diameterwas dependent on the assimilate supply after bud break. Axillarybuds obviously need a certain developmental stage to be ableto break. When released from correlative inhibition at an earlierstage, increased leaf initiation occurs before bud break.Copyright1994, 1999 Academic Press Age, axillary bud, cell number, cell size, pith, shoot growth, Rosa hybrida, rose     

Bud induction inSesamum indicum by splitting shoot apices followed by treatment with Amo-1618

Couples of buds were induced at the eccentric sites in the axial of the cotyledon inSesamum indicum by treating the embryo with a growth retardant Amo-1618 after the embyro shoot apex was split in the intercotyledonary plane, or incised between the primordia of the first opposite leaves. They appeared at first to have been transformed from the primordia of the first leaves. Developmental studies of the buds, however, revealed that they did not arise from the primordia, but from their adjacent area which is the presumptive stem tissue situated between the primordia and the cotyledon. Buds could occur only when the original shoot apex of the embryo as well as the first leaf primordia were degenerated. From experimental and circumstantial evidences, they were interpreted as axially buds induced at an unusual site.     

Micropropagation and field evaluation of micropropagated plants of turmeric

A protocol was developed for in vitro propagation of turmeric cv `elite' using young vegetative buds from sprouting rhizomes. The shoot buds produced multiple shoots when cultured on MS solid medium supplemented with benzyladenine and 1-naphthalene acetic acid. The effect of various cytokinins on shoot multiplication was studied by culturing the shoot tips on MS liquid medium supplemented with benzyladenine, benzyladenine riboside, kinetin, kinetin riboside, zeatin, 6-,-dimethylallylaminopurine, adenine, adenine sulfate or metatopolin each at 10 M in combination with 1-naphthalene acetic acid (1 M). Significant differences were observed between the treatments. Liquid medium was more favourable than agar medium for shoot multiplication. Among the various concentrations of agar tested, 0.4% and 0.6% were the best and produced the highest number of shoots per explant. Among the different carbohydrates tested, sucrose, fructose, glucose, sugar cubes, maltose, levulose and market sugar were found to be equally effective for shoot multiplication and xylose, rhamnose, lactose and soluble starch were inhibitory. Ninety five percent of the micropropagated plants survived in sterilized soil in paper cups and all of them survived in the field. Among 48 plants, two plants showed variegated leaves on the tillers. The micropropagated plants showed a significant increase in shoot length, number of tillers, number and length of leaves, number of fingers and total fresh rhizome weight per plant when compared with conventionally propagated plants. RAPD analysis of 11 regenerated plants using sixteen 10-mer primers did not show any polymorphism.     

Cytokinins in Rosa hybrida in relation to bud break

To assess the role of endogenous cytokinins in growth and development of Rosa hybrida , their concentrations in bleeding sap and in roots, stem, leaves, axillary shoots and bottom breaks in three stages of development were quantified. Cytokinins were purified by means of immunoaffinity chromatography and HPLC, and identified by retention time, UV spectrum and GC-MS. The major translocation form in the xylem was zeatin riboside (ZR). In all mature tissues, cytokinins of the zeatin-type were predominant, amounting to 80–90% of the total cytokinin concentration. The stems contained high concentrations of cytokinins, probably caused by lateral movement of ZR from the xylem to adjacent stem tissue and the ability of the stem to metabolize cytokinins. In young leaves the contribution of isopentenyl adenine (iP)-type cytokinins to the total cytokinin pool was about 50%, indicating that these leaves might be capable of de novo synthesis of cytokinins. In older leaves, the concentration of an unidentified cytokinin-like compound increased to more than 50% of total cytokinins. This compound, which was also found in the roots, might be a storage form of cytokinins. In young axillary shoots, about 50% of the cytokinins are iP-compounds, suggesting either import of iP-type cytokinins via the phloem or de novo synthesis of cytokinins. In buds forming bottom breaks, ZR and zeatin riboside monophosphate (ZRMP) are the main cytokinins, indicating that these buds receive their cytokinins from the roots.