Micropropagation of olive (<Emphasis Type="Italic">Olea europaea</Emphasis> L.) and application of mycorrhiza to improve plantlet establishment

Micropropagation methods were developed for the three French varieties of olive (Olea europaea L.), Aglandau and Tanche, that have the Appelation d’Origine Contrôlée (AOC) status and one ecotype (05300, Laragne, France). Explants consisting of partially lignified nodal segments were collected from rejuvenated glasshouse-grown plant material. Nodal explants with axillary buds were cultured on different media. For AOC varieties, olive medium modified (OM mod) to contain half the concentration of macroelements was the most efficient in inducing bud break and growth when supplemented with 30 g l^?1sucrose and 4 mg l^?1 zeatin. The resulting shoot buds were further multiplied and maintained on OM mod medium. Rooting was best achieved on OM supplemented with 4 mg l^?1 indole-3-butyric acid (IBA). For the Laragne ecotype, maximal shoot proliferation occurred when explants were cultured on woody plant medium supplemented with 15 g l^?1 sucrose and 0.1 mg l^?1 zeatin. Efficient rooting was achieved with 1 mg l^?1 IBA combined with 0.75 mg l^?1 naphthaleneacetic acid. After acclimatization in the glasshouse, survival rates ranged from 57 to 92%, depending on the genotype. Inoculation of Laragne ecotype microplantlets with the arbuscular mycorrhizal fungus Glomus mosseae significantly improved plant survival and subsequent plant development and growth.     

In vitro formation of axillary buds by immature shoots of Ponderosa pine

Axillary buds were induced from immature shoot explants taken from terminal buds of branches from 29- and 34-year old ponderosa pines (Pinus ponderosa Dougl ex Laws). The effect of collection time, position on the donor tree from which the explants were taken, and plant growth regulators on axillary bud formation was investigated. Explants from branches taken in late October formed axillary buds, whereas explants from branches collected in February 1988 produced a large amount of callus. The ability to form axillary buds was significantly greater for explants from the upper crown than from the lower portion of the tree. Explant elongation occurred and basal needle primordia swelled on Murashige & Skoog media (MS) containing 2.2 M 6-benzyladenine (BA) and 5.4 M naphthalenacetic acid. When transferred to a MS medium containing 4.4 M BA, 59% of explants formed axillary buds.     

红厚壳茎段丛生芽诱导与植株再生

红厚壳(Calophyllum inophyllum)为藤黄科红厚壳属多年生木本植物,有很高的药用价值。该研究以红厚壳带节茎段为外植体,探讨生长调节剂对腋芽萌发及丛生芽诱导、伸长和试管苗生根的影响。研究结果表明,外植体腋芽萌发和丛生芽诱导效果最好的培养基是MS+NAA1.0+TDZ0.5,在此条件下培养21天后,转入添加0.5 g·L–1活性炭且无生长调节剂的MS培养基,可有效促进不定芽的伸长。将带不定芽的外植体先在附加1.0 mg·L–1NAA的1/2MS培养基上进行生根诱导4周,之后转入附加1.0 g·L–1活性炭的无激素培养基进行根的伸长培养,这样的两步生根法能有效促进红厚壳生根。     

In vitro shoot regeneration from leaf explants of Roman Chamomile

Murashige and Skoog (1962) medium supplemented with 1.0 to 4.5 M of BA and 1.0 M of NAA induced adventitious bud formation and shoot development in leaf explants of Roman Chamomile. A higher number of adventitious buds was observed at the proximal end of the explants. Plantlets were replicated and multiplied on MS medium supplemented with 2.25 M of BA and 0.6 M of IAA. Plantlets were rooted on MS medium supplemented with 0.5 M of IBA and successfully weaned in vivo. The plants grew to maturity with high uniformity and no morphological signs of somaclonal variation.     

濒危植物盐桦离体组织培养特性的研究

目的:探讨新疆濒危植物盐桦离体组织培养的特性。方法:从盐桦原生地阿尔泰阿拉哈克盐湖边采摘盐桦休眠实生苗上的落叶枝条,待其萌发后分别取带芽嫩茎、嫩茎茎段及嫩叶芽尖三种不同材料接种于启动培养基,比较三种盐桦离体组织的诱导分化,继而设计不同激素、不同水平的单因子试验和正交试验,筛选适宜盐桦外植体芽增殖和生根的分化培养基。结果:诱导盐桦芽增殖的最佳外植体是带芽嫩茎,盐桦外植体增殖、壮苗最适培养基为:MS 6-BA 1.0mg/L IBA 0.5mg/L;盐桦外植体生根最适培养基为:1/2MS IBA 0.5mg/L 蔗糖30g/L 琼脂7% 暗光处理3d。结论:本研究筛选获得适宜盐桦芽增殖和生根的最佳培养条件,为高效扩繁和保存盐桦种质资源奠定了基础。     

Micropropagation of Eucalyptus tereticornis Smith.

Axillary shoot bud multiplication has been achieved in Eucalyptus tereticornis Smith. using explants from different regions of 8–10 years old elite trees, growing in the field. Results showed that addition of NAA at 0.1 mgl^-1 and BAP at 1.0 mgl^-1 to modified MS medium induced maximum number of shoot buds. For inducing axial growth in regenerated bud promordia, the hormone concentration of the medium was lowered. The addition of charcoal and gibberellic acid to the medium were beneficial. Rooting was best in Knop's medium containing 1.0 mgl^-1 IBA. The key factor in root induction was primarily a dark incubation for a short period. The percentage of both rooting of shoots and survival of the rooted shoots was 60–80.Continuous trials using explants from the elite trees throughout the year showed that the period between July–September was the best season for the explant source for rapid and increased multiplication of axillary buds. Phenolic exudation was also minimum at this period. The experiments were repeated using 50 populations from different plantations. It was observed that during culture, genotypically different populations responded differently in spite of optimal growth conditions.     

Direct shoot regeneration from leaf explants of Jatropha curcas in response to thidiazuron and high copper contents in the medium

Leaf explants of Jatropha curcas cultured on Murashige and Skoog’s (MS) medium supplemented with thidiazuron (TDZ; 0.90 μM) in combination with indole-3-butyric acid (IBA; 0.98μM) produced adventitious shoot buds directly on the surface of the explants without formation of intervening callus while shoot bud formation was accompanied with callus formation on medium supplemented with 6-benzylaminopurine (BAP; 13.3 μM) and IBA (2.46 μM). TDZ treatment resulted in more than twice higher rate of shoot bud induction than BAP. Shoot buds were multiplied and elongated following repeated transfers to medium containing BAP (2.22 μM) and gibberellic acid (GA₃; 1.44 μM). The effect of copper sulphate on differentiation of shoot buds from leaf segments was also investigated. Both shoot induction and multiplication media were supplemented with different levels of CuSO₄ (0–5 μM). Significant improvement in shoot bud induction was observed when the concentration of CuSO₄ was increased to 10 times the normal MS level. Healthy elongated shoots were rooted on half strength MS medium supplemented with IBA (2.46 μM). Rooted plantlets were transferred to field and survived. Histological analysis revealed direct formation of shoot buds from leaf explants.     

Micropropagation of wild beet (Beta maritima) from inflorescence pieces

In order to investigate the regeneration of wild beet (Beta maritima) from inflorescence pieces, the effects of growth regulator, genotype, explant source and stage of plant development on adventitious shoot formation and rooting in vitro and subsequent transplanting in the glasshouse were tested. Inflorescence tips produced more adventitious shoots than sub-apical segments and the best micropropagation was achieved on a Murashige and Skoog (MS) medium supplemented with 1.0 mg l^–1 BAP. Addition of auxin was not beneficial. The induction rate of adventitious shoots was genotype-dependent and influenced by the stage of plant development. Adventitious shoots were produced from the base of the flower buds, i.e. from the receptacle, not from axils or stalks and only a few buds on inflorescence tip explants produced adventitious shoots. Rooting was increased by using a MS medium with 3% sucrose supplemented with 1.0 mg l^–1 NAA. There was no variation in leaf morphology of the transplants. This work shows that inflorescence tips can be used successfully as explants for in vitro multiplication of sugar beet and wild beet.Abbreviations BAP benzylaminopurine - IBA indole-3-butyric acid - GA₃ gibberellic acid - MS Murashige and Skoog medium - NAA naphthaleneacetic acid Author for correspondence     

In vitro adventitious bud regeneration from internode segments of beech

Internode explants collected from in vitro grown shoots of two clones of Fagus sylvatica L. (European beech) and five clones of F. orientalis Lipski (Oriental beech) were used to evaluate their bud regeneration capacity. Adventitious shoot-buds formed on callus, which developed from internode segments cultured in a Woody Plant Medium supplemented with different concentrations of either thidiazuron (TDZ) or benzyladenine (BA). After 4 weeks of culture on induction media, the explants were transferred to a proliferation medium supplemented with 2.2 μM BA, 9.1 μM zeatin and 2.9 μM indole-3-acetic acid (IAA) for another 8 weeks. Medium containing TDZ was much more efficient than medium containing BA in inducing adventitious buds, the optimal TDZ concentration being 4.5 μM and the optimal BA concentration 17.8 μM. Genotypic variation in shoot regeneration capacity was observed among the two Fagus species and between clones within each species, with a significant interaction between TDZ concentration and genotype regarding mean bud number. Thidiazuron induction medium supplemented with a range of individual auxins was investigated, and it was found that IAA or indole-3-butyric acid at 2.9 μM enhanced the bud forming capacity of explants. Morphogenic response varied significantly with the position of the internode along the stem. The highest regeneration potential was obtained from apical internodes, while those distal to the apex were the least productive. Elongated shoots of adventitious origin can be readily proliferated by axillary branching. This revised version was published online in June 2006 with corrections to the Cover Date.     

马尾松高效再生体系的建立(简报)

利用组织培养技术快速繁殖针叶树,不仅为植树造林所需大量苗木开辟了一条快速高效的新途径,是生产优良无性系的捷径,而且也是植物基因工程技术应用于针叶树品种改良的前提条件。它将在林木种苗产业化和林木良种化进程中发挥重要作用。针叶树的离体培养和植株再生一直是植物组织培养研究中难度较大的一个领域。近几十年来,随着世界各国对发展无性系育林业的日益重视,针叶树组织培养研究取得了很大的进展[1-3]。马尾松(Pinus massoniana Lamb.)是我国南方主要的造林及绿化树种,其木材和松脂具有重要的经济价值。然而在生产实践中,种子园种子…     

<Emphasis Type="Italic">In vitro</Emphasis> bud regeneration of <Emphasis Type="Italic">Carthamus tinctorius</Emphasis> and wild <Emphasis Type="Italic">Carthamus</Emphasis> species from leaf explants and axillary buds

The organogenic competence of leaf explants of eleven Carthamus species including C. tinctorius on Murashige and Skoog (MS) medium supplemented with different concentrations of thidiazuron (TDZ) + α-naphthaleneacetic acid (NAA) and 6-benzyladenine (BA) + NAA was investigated. Highly prolific adventitious shoot regeneration was observed in C. tinctorius and C. arborescens on both growth regulator combinations and the shoot regeneration frequency was higher on medium supplemented with TDZ + NAA. Nodal culture of nine Carthamus species on media supplemented with BA and kinetin (KIN) individually revealed the superiority of media supplemented with BA over that of KIN in facilitating a higher shoot proliferation index. Proliferating shoots from axillary buds and leaf explants were transferred to medium supplemented with 1.0 mg dm⁻³ KIN or 0.5 mg dm⁻³ BA for shoot elongation. Elongated shoots were rooted on half-strength MS medium supplemented with 1.0 mg dm⁻³ each of indole-butyric acid (IBA) and phloroglucinol. The plantlets thus obtained were hardened and transferred to soil.     

Shoot regeneration of dwarf dogwood (Cornus canadensis L.) and morphological characterization of the regenerated plants

Dwarf dogwoods (or the bunchberries) are the only suffrutex in Cornaceae. They are attractive ground cover ornamentals with clusters of small flowers surrounded by petaloid bracts. Little has been reported on plant regeneration of dogwoods. As a step toward unraveling the molecular basis of inflorescence evolution in Cornus, we report an efficient regeneration system for a dwarf dogwood species C. canadensis through organogenesis from rejuvenated leaves, and characterize the development of the plantlets. We used the nodal stem segments of vegetative branches as explants. Micropropogated shoots were quickly induced from axillary buds of nodes on an induction medium consisting of basal MS medium supplemented with 4.44 μM BAP and 0.54 μM NAA. The new leaves of adventitious shoots were used as explants to induce calli on the same induction medium. Nearly 65% of leaf explants produced calli, 80% of which formed adventitious buds. Gibberellic acid (1.45 μM) added to the same induction medium efficiently promoted quick elongation of most adventitious buds, and 0.49 μM IBA added to the basal MS medium promoted root formation from nearly 50% of the elongated shoots. The growth of plantlets in pot soil was characterized by the development of functional woody rhizomes, which continuously developed new aboveground vegetative branches, but not flowering branches, within the past 12 months. Potential reasons causing the delay of flowering of the regenerated plants are discussed. The establishment of this regeneration system facilitates developing a genetic transformation system to test candidate genes involved in the developmental divergence of inflorescences in Cornus. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

大果良种沙棘愈伤组织诱导及植株再生的研究

大果良种沙棘的幼嫩茎尖,茎段外植体接种在MS,1／2MS附加不同浓度配比的IAA,IBA,BA,NAA培养基上可诱导茎尖及腋芽生长,将诱导产生的无性系芽接种在MS或1／2MS附加BA0．3－0．5mg/L,NAA0．05mg/L的培养基上可形成丛生芽,同时在小叶片和嫩茎上诱导产生愈伤组织,继续培养愈伤组织表面形成大量的绿色突起,进一步分化成不定芽,在相同培养基上,不定芽上可直接产生不定芽,从而形成多达数百个的不定芽族,不定芽长至3cm时切下转至1／2MS附加IAA或IBA 0．2mg/L的培养基上可生根而形成完整 的再生植株。     

Plantlet regeneration from mature embryos of Juniperus cedrus

A protocol is described for plantlet regeneration using embryonic explants of Juniperus cedrus Webb & Berth. An average of 6 adventitious buds were induced on whole excised embryos cultured for 15 days on Quoirin and LePoivre (QP) half-strength medium supplemented with 5 M N⁶-benzyladenine. For bud development, explants were transferred to phytohormone-free 1/2 QP medium. For shoot elongation, explants were cultured on 1/2 QP with 0.05% activated charcoal and 2% sucrose. Adventitious shoots were rooted successfully in peat-vermiculite-perlite (1:1:1) moistened with 1/4 QP containing 1% sucrose and 5 M -napthaleneacetic acid, pH 5.0. Axillary shoots elongated spontaneously in culture from leaf axils.Abbreviations AC activated charcoal - BA N⁶-benzyladenine - NAA -naphthaleneacetic acid - QP Quoirin & LePoivre - SH Schenk & Hildebrandt     

铁核桃离体培养与快速繁殖

以优良单株‘纳雍-1’的单芽茎段为外植体,建立了铁核桃（Juglanssigillata）离体培养与快速繁殖的体系。结果表明,附加6-BA1．0mg·L-1 ＋活·IgK（AC）3．0g·L-1的DKw培养基适宜铁核桃腋芽诱导;适宜铁核桃芽增殖的培养基为DKW＋6-BA1．0mg·L-1 ＋IBA0．02mg·L-1,40d后增殖系数可达7．33;试管苗的茎尖和茎段均可用于增殖培养;一步生根法（低浓度的生长素IBA持续诱导）不利于铁核桃试管苗嫩茎生根;采用二步生根法,生根率最高可达71．73％,其中,不同IBA浓度、暗培养时间、蔗糖浓度和AC含量对试管苗嫩茎生根影响显著,铁核桃试管苗在附]sulBA5,0mg·L-1的1／4DKW培养基中暗培养12d,再转移到不含IBA的1／4DKW培养基（附加AC 3g-L-1和蔗糖20g·L-1）中生根效果最好;生根试管苗采用珍珠岩和营养土两步炼苗,60d后成活率达到87．50％。     

Regeneration of plants from leaflet explants of tissue culture raised safed siris (Albizia procera)

Shoot bud regeneration was obtained from isolated leaflets of Albizia procera cultured on MS medium with various concentrations of 6-benzyladenine (BA) and α-naphthaleneacetic acid (NAA). The highest numbers of adventitious buds were obtained on MS medium supplemented with 10 μM BA and 1 μM NAA. The replacement of 7 g l-1 Difco bacto agar with 2.6 g l-1 Phytagel in the medium enhanced adventitious bud regeneration. Further, addition of 15 μM silver nitrate promoted callus-free shoot regeneration from leaf explants. The regenerated shoot buds were elongated on MS medium containing 0.01 μM BA and 1 μM NAA. Rooting was obtained on modified MS medium supplemented with 2 μM IBA. To our knowledge this is the first report of direct regeneration of shoots from leaflet explants in A. procera, and should help facilitate genetic transformation in this species. This revised version was published online in June 2006 with corrections to the Cover Date.     

Optimization of in vitro bud induction and plantlet formation from mature embryos of Aleppo pine (Pinus halepensis Mill.)

Summary Studies were undertaken to optimize tissue culture conditions for micropropagation of Aleppo pine (Pinus halepensis Mill.) from mature embryos and various explants of the embryo. Over 90% of the embryo explants gave rise to adventitious buds within 4 wk. Intact embryos were the most suitable explants for shoot bud induction. Both isolated cotyledons and hypocotyls produced adventitious buds, but these developed slowly and failed to elongate. N⁶-Benzyladenine (BA) alone at 5.0μM was the most effective cytokinin when added to gelled to gelled von Arnold and Eriksson’s (AE) medium containing 3% sucrose. Adventitious bud development was achieved on hormone-free AE medium, and shoot elongation was optimum on three quarter-strength Bornman’s MCM medium, with 0.1% conifer-derived activated charcoal. Shoots were multiplied on three-quarter strength MCM medium, containing 5μM BA. To induce adventitious roots on the elongated shoots, pulse treatment with 1 mM IBA for 6 h, followed by the transfer of the shoots to sterile peat:vermiculite (1:1) mixture, was beneficial. After acclimatization for 3 to 4 wk under mist, almost all the rooted shoots could be transplanted successfully to the greenhouse, where the plants exhibited normal growth habit. Histologic studies on the ontogeny of adventitious shoot formation from mature embryo explants revealed temporal structural changes in different parts of the explant. Induction of mitotic divisions on the shoot-forming medium resulted in the formation of meristemoids in the epidermal and subepidermal layers of the explant, located initially at both the tips of the cotyledons and the axils of adjacent cotyledons. Shoot buds arising in the axils of adjacent cotyledons were due to new cell division and not to any preexisting meristem.     

Plant regeneration via adventitious bud formation from cotyledon explants of Panax ginseng C. A. Meyer

Cotyledon explants of ginseng (Panax ginseng C. A. Meyer) produced somatic embryos directly on medium without growth regulators, with 89% of the explants forming somatic embryos. Cytokinin treatment greatly suppressed somatic embryo formation but stimulated the direct formation of adventitious buds. BAP treatment was more effective than the kinetin treatment for adventitious bud formation. Auxin (0.05 mg/l IBA) in combination with cytokinin enhanced adventitious bud formation, with the highest frequency, 40%, at 0.05 mg/l IBA and 5 mg/l BAP. Adventitious buds were mainly formed near the distal portion of the cotyledons, while somatic embryos were formed near the proximal excised margins. Shoots were developed from adventitious buds after transfer to MS medium with 10 mg/l GA₃. Root formation from the shoots was obtained after the shoots were transferred to half-strength MS medium with auxin (IAA). When the plants derived from adventitious buds were transferred to greenhouse soil, 36% were successfully acclimatized. Received: 7 November 1997 / Revision received: 12 January 1998 / Accepted: 7 February 1998     

Relationship of embryogenic competence in maize (Zea mays L.) leaves to mitotic activity,cell cycle and nuclear DNA content

Axillary bud explants of 11 selected mature waratah clones were established in vitro on a modified Murashige & Skoog medium. Adequate proliferation of axillary shoots was achieved by optimisation of the growth regulator status of the culture medium. For the majority of clones, a three to six times rate of proliferation was achieved with 1.25 M BA and 1.0 M GA₃ without the occurrence of abnormalities. The white flowering clone did not respond favourably to the addition of GA₃ to the medium.Abbreviations BA benzyladenine - GA₃ gibberellic acid - IBA indole-3-butyric acid - LSD least significant difference - MS Murashige & Skoog medium     

Micropropagation of Alnus cordata (Loisel.) Loisel.

Procedures were developed for micropropagation of Alnus cordata through in vitro axillary shoot multiplication of axillary bud explants cultured in Murashige & Skoog (MS) medium. Establishment of cultures from plants grown in the field was very difficult due to bacterial contamination and phenolic oxidation in explants causing severe browning. Explants were first cultured on an MS medium containing 4.4 M 6-benzyladenine and 87.6 mM sucrose (initiation medium) for 7 days and then transferred to an MS medium containing 1.1 M 6-benzyladenine and 333 mM glucose (multiplication medium) for a further 20–25 days. It was necessary to transfer cultures from initiation medium to multiplication medium after 7 days to minimize excessive callus growth, abnormally thick and brittle leaves, inhibition of shoot elongation, and senescence. Shoot multiplication comparable to the above method was achieved by culture of axillary bud explants in MS medium supplemented with 1.1–4.4 M 6-benzyladenine and 333 mM glucose 4–5 weeks after culture. Shoots rooted in MS medium (1/2 x macro-nutrients) supplemented with 1.2–4.9 M indolebutyric acid. Also, 98% rooting was achieved when cultures were treated with 625 mgl^-1 indolebutyric acid for 24 h at the end of the shoot production stage and rooted in vivo as mini-cuttings. Plantlets established well in soil.