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1.
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Three explants namely, nodal, internodal and petiolar segments were used to establish in vitro cultures of Piper longum. Multiple shoots were induced on semi-solid Murashige and Skoog (MS) medium supplemented with 1 mg/l 6-benzyladenine (BA).
Addition of ascorbic acid (40 mg/l) considerably reduced browning of tissue and medium. Best shoot regeneration was observed
from petiolar explants and was, therefore, used for all further studies. An indexing method was introduced for checking bacterial
contamination in well established shoot multiplication cultures. It was found that bacterial infection was quite high in shoots
derived from nodal and internodal explants while it was least in those obtained from petiolar segments. Only shoots that indexed
negative for endogenous bacteria were used for proliferation and in vitro conservation studies. At the end of 4 weeks in proliferation
medium which consisted of MS supplemented with 0.5 mg/l BA and 40 mg/l ascorbic acid as many as 22 shoot buds of 41 mm length
could be obtained. Shoot buds developed into clusters for ease of further proliferation. A step of shoot elongation for 2 weeks
in liquid MS basal medium was found to be beneficial for getting long and healthy shoots for rooting. Single shoots were rooted
in 0.25 mg/l indole butyric acid that could be successfully acclimatized under nethouse conditions. A conservation strategy
was also developed. The shoot cultures could be maintained without subculturing for as long as 8 weeks in MS medium supplemented
with 1 mg/l paclobutrazol (PBZ) and 40 mg/l ascorbic acid. 相似文献
3.
猕猴桃高频直接再生体系的建立 总被引:3,自引:0,他引:3
为了建立猕猴桃高频再生体系,以MS为基本培养基,猕猴桃(Actinidia deliciosaQinmei)茎及叶片为外植体,研究了2,4-D、6-BA和NAA在美味猕猴桃愈伤组织形成及分化过程中的作用。方差分析结果表明,6-BA能够显著促进愈伤组织形成,6-BA和NAA可以显著促进愈伤组织形成和分化,而2,4-D抑制愈伤组织形成。附加2.0 mg/L 6-BA、1.0 mg/L NAA和600 mg/L水解酪蛋白的MS培养基是茎段培养的最佳培养基,在该培养基上,以再生的无菌苗为起始材料,一个月时叶圆盘的直接再生频率达到100%,平均每个叶圆盘产生9.33个芽,其中23.21%芽高度超过0.5 cm。 相似文献
4.
Wongwicha W Tanaka H Shoyama Y Tuvshintogtokh I Putalun W 《Zeitschrift für Naturforschung. C, Journal of biosciences》2008,63(5-6):413-417
Licorice plants, Glycyrrhiza glabra, G. uralensis, and G. inflata, were investigated for callus induction using Murashige and Skoog (MS) medium combined with auxins and cytokinins. After 4 weeks of culture, 33-100% of leaf or stem explants formed calli. Maximum of shoot induction from callus cultures was achieved by G. inflata stem explants cultured on MS medium supplemented with 1 mg/l alpha-naphthaleneacetic acid (NAA) and 0.5 mg/l 6-benzyladenine (BA) (67%) which also gave maximum shoot formation per explant (two shoots per explant). These results indicated that all three Glycyrrhiza species regenerated shoots from callus cultures on MS medium combined with NAA and BA or only thidiazuron (TDZ; 0.1 and 0.5 mg/l). Glycyrrhizin contents of G. uralensis calli induced using MS medium in combination with NAA and BA [(27.60 +/- 8.47) microg/g DW] or TDZ alone [(36.52 +/- 2.45) microg/ g DW] were higher than those found in other combinations. 相似文献
5.
Yaser Hassan Dewir Nisha Singh Shakira Shaik Ashley Nicholas 《In vitro cellular & developmental biology. Plant》2010,46(1):41-46
The present study reports a simple protocol for indirect shoot organogenesis and plant regeneration of Sutherlandia using rachis and stem segments. Different concentrations (0.0–68.08 μmol l−1) of thidiazuron (TDZ) were used for callus induction and shoot organogenesis. The highest percentage of callus formation
(97.5%) and the highest percentage of explants forming shoots (88.8%) were obtained from rachis explants cultured onto Murashige
and Skoog (MS) medium (Murashige and Skoog, Physiol. Plant. 15:473–495, 1962) supplemented with 45.41 μmol l−1 TDZ. Scanning electron microscopy demonstrated the early development of adventitious shoots derived from callus cultures.
Shoot clusters were further developed and grown in MS hormone-free medium. The presence of l-canavanine was determined by thin-layer chromatography and confirmed after column fractionation using silica gel and nuclear
magnetic resonance spectroscopy. Individual shoots were rooted on different concentrations and combinations of MS salt strength
and IBA. Half-strength MS salt medium supplemented with 24.6 μmol l−1 IBA was optimal for root induction in which 78% of shoots were rooted. The in vitro plants were successfully acclimatized in a growth chamber with a 90% survival rate. 相似文献
6.
Summary A method has been developed for the induction of adventitious shoots from leaf tissue of Echinacea pallida with subsequent whole-plant regeneration. Proliferating callus and shoot cultures were derived from leaf tissue explants
placed on Murashige and Skoog medium supplemented with 6-benzylaminopurine and naphthaleneacetic acid combinations. The optimum
shoot regeneration frequency (63%) and number of shoots per explant (2.3 shoots per explant) was achieved using media supplemented
with 26.6 μM 6-benzylaminopurine and 0.11 μM naphthaleneacetic acid. Rooting of regenerated shoot explants was successful on Murashige and Skoog medium, both with and
without the addition of indole-3-butyric acid. All plantlets survived acclimatization, producing phenotypically normal plants
in the greenhouse. This study demonstrates that leaf tissue of E. pallida is competent for adventitious shoot regeneration and establishes a useful method for the micropropagation of this important
medicinal plant. 相似文献
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8.
Janhvi Mishra Himani Bhandari Mahendra Singh Sandeep Rawat R. K. Agnihotri S. Mishra S. Purohit 《Acta Physiologiae Plantarum》2011,33(5):1841-1846
Picrorhiza kurroa Royle ex Benth. is an endangered plant producing various compounds of medicinal importance. Hairy roots of P. kurroa were obtained following cocultivation of shoot tip explants with Agrobacterium rhizogenes strains A 4 and PAT 405. Bacterial strain A 4 appeared to be better than the strain PAT 405 in terms of both growth of respective
hairy root cultures and secondary metabolite production. The optimal growth of both the hairy root cultures occurred on half-strength
semisolid medium with 3% sucrose. Picrotin and picrotoxinin from the roots of wild type field grown plants were compared with
8-week-old hairy root cultures induced by the A 4 and PAT 405 strains of A. rhizogenes. Picrotin and picrotoxinin content were evaluated in hairy root cultures as well as roots of field grown plant of P. kurroa. In terms of the production of picrotin and picrotoxinin, the A 4 induced hairy roots appeared to be a better performer than
the PAT 405 induced hairy root cultures. The picrotin and picrotoxinin content was highest in 8-week-old A 4 induced hairy
roots (8.8 μg/g DW and 47.1 μg/g DW, respectively). Rapid growth of the hairy roots of P. kurroa with in vitro secondary metabolite production potential may offer an attractive alternative to the exploitation of this endangered
plant species. 相似文献
9.
Boon Chin Tan Chiew Foan Chin Peter Alderson 《Plant Cell, Tissue and Organ Culture》2011,105(3):457-463
An indirect in vitro plant regeneration protocol for Vanilla planifolia has been established. Juvenile leaf and nodal segments from V. planifolia were used as explants to initiate callus. Nodal explants showed better callus initiation than juvenile leaf explants, with
35.0% of explants forming callus when cultured on Murashige and Skoog (MS) basal medium supplemented with 2.0 mg/l 1-naphthylacetic
acid (NAA) and 1.0 mg/l 6-benzyladenine (BA). Almost 10.0% of juvenile leaf explants were induced to form callus on the MS
basal medium containing 2.0 mg/l NAA and 2.0 mg/l BA, whereas no callus formed in the presence of any concentrations of 2,4-dichlorophenoxyacetic
acid (2,4-D) and BA. After 8 weeks, callus generated was transferred to MS basal medium containing 1.0 mg/l BA and 0.5 mg/l
NAA. A mean number of 4.2 shoots per callus was produced on this medium, with a mean length of 3.8 cm after 8 weeks of culture.
Roots formed on 88.3% of plantlets when they were cultured on MS medium supplemented with 1.0 mg/l NAA, with a mean length
of 4.4 cm after 4 weeks of culture. Of the rooted plantlets, 90.0% survived acclimatisation and were making new growth after
4 weeks. 相似文献
10.
Explants of shoot tips, internodal stem sections, and leaf segments of Lisianthus, Eustoma grandiflorum (Griseb.) Schinners, Dwarf Purple were cultured in vitro on modified Murashige and Skoog (MS) media. Explants of shoot tips and internodal stem sections developed into multiple shoots, whereas, leaf segments turned chlorotic on a medium supplemented with 3 mgl-1 benzyladenine (BA) and 0.2 mgl-1 naphthalene acetic acid (NAA). Shoot proliferation was obtained on shoot tips and leaf segments with 3 mgl-1 BA, but internodal stem sections became necrotic and died on this medium. Rooting was induced in cultures with multiple shoots by subculturing explants on a half-strength MS medium supplemented with 2 mgl-1 indole-3-acetic acid (IAA). Rooted plantlets were successfully transferred to soil. 相似文献
11.
Bimal Kumar Ghimire Chang Yeon Yu Ill-Min Chung 《Plant Cell, Tissue and Organ Culture》2012,108(3):455-464
A simple and efficient procedure was developed for in vitro propagation of Solanum aculeatissimum Jacq. using leaf and petiole explants cultured on Murashige and Skoog (MS) medium supplemented with α-naphthalene acetic
acid (NAA) and 6-benzyladenine (BA). Effects of various plant growth regulators, explant types, carbohydrates, and basal salts
on induction of adventitious shoots were also studied. Leaf explants appeared to have better regeneration capacity than petiole
explants in the tested media. The highest regeneration frequency (79.33 ± 3.60%) and shoot number (11.33 ± 2.21 shoots per
explant) were obtained in leaf explants in MS medium containing 3% sucrose and 0.8% agar, supplemented with 0.1 mg/l NAA and
2.0 mg/l BA, whereas petiole explants were more responsive to 0.1 mg/l NAA and 1.0 mg/l thiadiazuron. Developed shoots rooted
best on MS medium with 1.0 mg/l indole acetic acid (IAA), producing 18.33 ± 2.51 roots per shoot. Histological investigation
showed that the shoot buds originated mainly from epidermal cells of wounded tissues, without callus formation. The regenerated
plantlets were successfully acclimatized in a greenhouse, where over 90% developed into morphologically normal and fertile
plants. Results of flow cytometry analysis on S. aculeatissimum indicated no variation in the ploidy levels of plants regenerated via direct shoot formation and showed almost the same phenotype
as that of mother plants. This adventitious shoot regeneration method may be used for large-scale shoot propagation and genetic
engineering studies of S. aculeatissimum. 相似文献
12.
Xiuli Shen Michael E. Kane Jianjun Chen 《In vitro cellular & developmental biology. Plant》2008,44(4):282-288
The capacity for indirect shoot organogenesis of leaf and root explants of four Dieffenbachia cultivars were examined on a modified Murashige and Skoog (MS; Physiol Plant 15:473–495, 1962) medium supplemented with different plant growth regulators in 112 combinations. Callus formation was only observed from
leaf explants on MS supplemented with 1–10 μM thidiazuron (TDZ) and 0.5–1.0 μM 2,4-dichlorophenoxyacetic acid (2,4-D) regardless
of cultivars. The combination of 5 μM TDZ and 1 μM 2,4-D resulted in the greatest callus formation frequency among the four
cultivars tested. Significant differences in callus and shoot formation from leaf explants were also observed among cultivars.
Cultivars Camouflage, Camille, Octopus, and Star Bright produced green nodular, brown nodular, yellow friable, and green compact
calli with corresponding maximum callus formation frequencies of 96%, 62%, 54%, and 52%, respectively. A maximum of 6.7 shoots/callus
was observed in cv. Camouflage, followed by cvs. Camille and Star Bright at 3.7 and 3.5, respectively. Calli of cv. Octopus
displayed no capacity for shoot organogenesis. Regardless of cultivar, callus formation was not observed on root explants.
Regenerated shoots were successfully acclimatized in a shaded greenhouse condition with 100% survival. 相似文献
13.
The effect of various hormonal combinations on callus formation and regeneration of shoot and root from leaf derived callus
of Acanthophyllum sordidum Bunge ex Boiss. has been studied. Proteins and activity of antioxidant enzymes were also evaluated during shoot and root organogenesis
from callus. Calli were induced from leaf explants excised from 30-d-old seedlings grown on Murashige and Skoog medium containing
4.52 μM 2,4-dichlorophenoxyacetic acid + 4.65 μM kinetin. Maximum growth of calli and the most efficient regeneration of shoots
and roots occurred with 2.69 μM 1-naphthalene acetic acid (NAA), 2.69 μM NAA + 4.54 μM thidiazuron and 2.46 μM indole-3-butyric
acid. Protein content decreased in calli and increased significantly during regeneration of shoots from callus. Superoxide
dismutase activity decreased in calli comparing to that of seedlings, then increased in regenerated shoots and roots. High
catalase activity was detected in seedlings and regenerated shoots, whereas high peroxidase activity was observed in calli
and regenerated roots. 相似文献
14.
Arzu Ucar Turker Esra Cansever Mutlu Arzu Birinci Yıldırım 《Acta Physiologiae Plantarum》2008,30(4):421-426
Epilobium angustifolium L. (fireweed) is a medicinal plant that has been used to treat diarrhea, mucous colitis, irritable-bowel syndrome, skin problems,
prostate problems, menstrual disorders, asthma, whooping cough, and hiccups. A highly efficient and rapid regeneration system
via multiple shoot formation was developed for fireweed. Explants (leaf, petiole, root, and stem segments) excised from sterile
seedlings were cultured on medium supplemented with different concentrations and combinations of various plant growth regulators.
Explant browning, a major problem for regeneration, was overcome by adding 100 mg/l ascorbic acid to all prepared media containing
growth regulator combinations. Root explants formed more shoots than other explants. Best shoot proliferation was obtained
from root explants cultured on media with 0.1 mg/l BA and 0.5 mg/l IAA. Regenerated shoots were transferred to rooting media
containing different concentrations of IAA, IBA, NAA or 2,4-D. Most shoots developed roots on medium with 0.5 mg/l IAA. Rooted
explants were transferred to vermiculate in Magenta containers for acclimatization and after 3 weeks they were planted in
to plastic pots containing potting soil and maintained in the plant growth room. 相似文献
15.
Kalina Danova Eva Čellárová Anna Macková Zuzana Daxnerová Veneta Kapchina-Toteva 《In vitro cellular & developmental biology. Plant》2010,46(5):422-429
The Balkan endemic species, Hypericum rumeliacum, Guttiferae was introduced in vitro for the first time with the aim to study the type of morphogenetic response to plant growth regulators and ability to produce
phenolics and flavonoid compounds. The morphoregulatory effect of 2,4-dichlorophenoxyacetic acid (2,4-D), 1-naphtaleneacetic
acid (NAA), 6-benzyladenine (BA) and combination of BA with NAA in Murashige–Skoog's basal medium on leaf lamina, internode
stem segment, stem node and root cuttings was studied. Histological analysis of the structures regenerated from the primary
explants proved the presence of both, embryoids and meristemoids. The node explants cultivated on BA-supplemented medium were
the most favourable for regeneration through meristemoids. Therefore a double-stage culture approach, allowing an effective
multiplication of large quantities of plant shoots in vitro along with maintenance of the biosynthetic capacity of the culture was developed. It comprised one subculture of three-nodal
stem explants derived from the stock shoot cultures on MS medium supplemented with 0.2 mg/l BA followed by subculture of the
induced multiple shoots on cytokinin-free MS medium. Determination of the total phenolics and flavonoids showed that the decrease
of the levels of these secondary metabolites is transitional, as the exclusionof BA from the medium resulted in an increase
of their total content. 相似文献
16.
Summary A viable protocol has been developed for direct and indirect shoot regeneration of Vernonia cinerea. To establish a stable and high-frequency plant regeneration system, leaf and stem explants were tested with different combinations
of α-naphthalene acetic acid (NAA), indole-3-acetic acid (IAA), and benzylaminopurine (BA). Lateral buds on nodal explants
grew into shoots within 2 wk of culture in Murashige and Skoog (MS) basal medium supplemented with 20.9 μM BA. Excision and culture of nodal segments from in vitro-raised shoots on fresh medium with the same concentration of BA facilitated development of more than 15 shoots per node.
Similarly leaf, nodal, and internodal explants were cultured on MS basal medium supplemented with different concentrations
of BA, NAA, and IAA either alone or in combinations for callus induction and organogenesis. Shoot buds and/or roots were regenerated
on callus. Shoot buds formed multiple shoots within 4 wk after incubation in induction medium. Adventitious buds and shoots
proliferated when callus was cut into pieces and subcultured on MS basal medium containing 20.9 μM BA and 5.3 μM NAA. This combination proved to be the best medium for enhanced adventitious shoot bud multiplication, generating a maximum
of 50 shoots in 4 wk. This medium was also used successfully for shoot proliferation in liquid medium. Root formation was
observed from callus induced in medium containing 8.05–13.4 μM NAA. Regenerated shoots exhibited flowering and root formation in MS basal medium without any growth regulators. Plantlets
established in the field showed 85% survival and exhibited identical morphological characteristics as the donor plant. 相似文献
17.
Anna Piovan Rosy Caniato Elsa Mariella Cappelletti Raffaella Filippini 《Plant Cell, Tissue and Organ Culture》2010,100(3):309-315
Efficient plant regeneration systems both from shoot segments and via callus organogenesis were developed for Kosteletzkya pentacarpos (L.) Ledeb., a rare and endangered Eurasian species. In the experiments with existing meristems, factors affecting shoot
proliferation, including explant type, i.e. decapitated and intact shoots, and plant growth regulators, indole-3-acetic acid
or kinetin, were investigated. Shoot proliferation was significantly affected by the type of explant, the hormones and their
interaction. The highest shoot multiplication rate was obtained from decapitated shoots. Increasing kinetin concentration
promoted shoot elongation regardless of explant type. In intact shoots, shoot length was also affected by increasing auxin
concentration, although this effect tends to decrease with higher concentration. Decapitated shoots were not responsive to
the addition of auxin. Micropropagation through organogenesis from callus was also investigated. Calli were obtained from
leaf, stem internode and root explants. Only the leaf-derived calli produced shoots and indole-3-acetic acid favoured increased
numbers of shoots. A number of experiments were conducted for rooting of in vitro produced shoots. All of them induced high
rooting frequency, the number and the length of roots being dependent on the strength of the basal medium. The use of 1–2 mg l−1 indole-3-butyric acid resulted in refining the optimal concentration for root elongation. The regenerated plants (70%) survived
and flowered in their first vegetative period. 相似文献
18.
Giovanni Iapichino Tony H. H. Chen Leslie H. Fuchigami 《Plant Cell, Tissue and Organ Culture》1991,27(1):37-43
The influence of the source of plant material (greenhouse-grown plants or in vitro shoot cultures), the type of tissue explant (shoot-tip, single-node stem segment, whole leaf, leaf strip or half-leaf section) and growth regulator concentration on shoot regeneration from somatic tissue of Rhododendron laetum × aurigeranum was evaluated. No regeneration response was obtained on explants from greenhouse-grown plants. Adventitious shoots were obtained from callus produced at the basal end of shoot-tip and single-node stem segment explants derived from in vitro-grown shoots cultured on Anderson's medium supplemented with 22.8 M IAA and 73.8 M 2iP. The greatest percentage of adventitious shoot regeneration (77%) was induced on leaf sections cultured in the presence of 22.8 M IAA and 147.6 M 2iP. Plant regeneration was accomplished with minimal callus formation. This technique represents a further step toward gene manipulation of Rhododendron.Abbreviations IAA
1-H-Indole-3-acetic acid
- 2iP
N-(3-methyl-2-Butenyl)-1H-purin-6 amine 相似文献
19.
Rubén Mallón Juan Rodríguez-Oubiña María Luz González 《Plant Cell, Tissue and Organ Culture》2011,106(3):523-530
In vitro culture is currently used to produce plant material for ex situ conservation of endangered species. In this study,
an efficient protocol for shoot regeneration from leaves and roots was developed for Centaurea ultreiae, a critically endangered species. Organogenesis from leaf and root explants was promoted by incubating these explants on
half-strength Murashige and Skoog (MS) medium in the presence of one of four different cytokinins [6-benzyladenine (BA), zeatin,
kinetin or N6-(2-isopentenyl) adenine (2iP)], each provided at five different levels. Shoot organogenesis was induced in both explants.
The best response, 90% of leaf explants producing a mean of 2.48 shoots per explants and 94.3% of root explants producing
a mean of 5.60 viable shoots per explants, was observed when explants were incubated on a medium containing 0.55 μM BA. Histological
studies revealed connectivity between vascular tissues of regenerated shoots and cambial cells of leaf explants. Moreover,
adventitious shoots were derived from pericycle cells of root explants and parenchymatic cells of callus tissues. 相似文献
20.
Qi Zhu Fengtao Wu Feng Ding Dong Ye Yongqin Chen Yi Li Yang Zhifan 《Plant Cell, Tissue and Organ Culture》2009,96(3):317-324
Dioscorea zingiberensis Wright has been cultivated as a pharmaceutical crop for production of diosgenin, a precursor for synthesis of various important
steroid drugs. Because breeding of D. zingiberensis through sexual hybridization is difficult due to its unstable sexuality and differences in timing of flowering in male and
female plants, gene transfer approaches may play a vital role in its genetic improvement. In this study, the Agrobacterium tumefaciens-mediated transformation of D. zingiberensis was investigated with leaves and calli as explants. The results showed that both leaf segments and callus pieces were sensitive
to 30 mg/l hygromycin and 50–60 mg/l kanamycin, and using calli as explants and addition of acetosyringone (AS) in cocultivation
medium were crucial for successful transformation. We first immersed callus explants in A. tumefaciens cells for 30 min and then transferred the explants onto a co-cultivation medium supplemented with 200 μM AS for 3 days. Three
days after, we cultured the infected explants on a selective medium containing 50 mg/l kanamycin and 100 mg/l timentin for
formation of kanamycin-resistant calli. After the kanamycin-resistant calli were produced, we transferred them onto fresh
selective medium for shoot induction. Finally, the kanamycin resistant shoots were rooted and the stable incorporation of
the transgene into the genome of D. zingiberensis plants was confirmed by GUS histochemical assay, PCR and Southern blot analyses. The method reported here can be used to
produce transgenic D. zingiberensis plants in 5 months and the transformation frequency is 24.8% based on the numbers of independent transgenic plants regenerated
from initial infected callus explants. 相似文献