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1.
We developed an efficient plant regeneration system from protoplasts for poplar (Populus alba L.). Protoplasts were isolated from 4-day-old suspension cultures derived from seed-induced calli with a yield of 6.96× 106 cells/g fresh weight cells and then cultured at a concentration of 2.5×105 cells/ml in NH4NO3-free Murashige and Skoog (MS) medium supplemented with 5 μM 2,4-dichlorophenoxyacetic acid (2,4-D), 0.05 μM thidiazuron (TDZ) and 0.5 M glucose as a osmoticum. The plating efficiency of the cultured protoplasts was calculated at 26.5% at day 7 and 31.7% at
day 14. Cell colonies were observed after culturing for 4 weeks. Regenerated colonies were propagated through subculture in
liquid MS medium supplemented with 5 μM 2,4-D. Buds were induced from regenerated calli on MS medium containing 10 μM kinetin or 1 μM TDZ. Regenerated shoots were rooted on half-strength MS medium, and the plantlets were transplanted in soil. Randomly amplified
polymorphic DNA analysis did not detect any DNA polymorphism among the regenerated plants.
Received: 7 March 1997 / Revision received: 16 June 1997 / Accepted: 5 July 1997 相似文献
2.
J. Ponsamuel D. V. Huhman B. G. Cassidy D. Post-Beittenmiller 《Plant cell reports》1998,17(5):373-378
Shoot buds were induced from plumular explants of peanut (Arachis hypogaea L., cv `Okrun') preconditioned on medium containing 2,4-dichlorophenoxyacetic acid and kinetin and then transferred to regeneration
medium containing benzylaminopurine and β-naphthoxyacetic acid. Buds differentiated 25 days following transfer to regeneration medium. Each explant produced 30 to
40 buds, but only 4 shoots. The remaining buds were dormant and did not produce shoots when maintained on regeneration medium.
Shoots were regenerated continuously, however, when explants were subsequently transferred to shoot conversion medium containing
1 μM brassin, benzylaminopurine and β-naphthoxyacetic acid, respectively. Approximately 5 shoots were harvested every 30 days after transfer to shoot conversion
medium for up to 7 months. No further shoot production was observed from explants maintained on regeneration medium without
brassin. Regenerated shoots could be rooted and produced viable seeds. This procedure provides an efficient and reliable system
for regeneration and transformation studies using cv `Okrun'.
Received: 9 April 1997 / Revision received: 27 August 1997 / Accepted: 20 September 1997 相似文献
3.
Emilio Maruyama Katsuaki Ishii Isao Kinoshita Kihachiro Ohba Akira Saito 《In vitro cellular & developmental biology. Plant》1997,33(2):131-135
Summary Shoot organogenesis of Guazuma crinita Mart. from root and petiole explants was obtained via adventitious bud formation. Root segments and petiole explants excised
from in vitro generated plantlets were cultured on woody plant medium (WPM) supplemented with [trans-6-(4-hydroxy-3-methylbut-2- enyl)aminopurine]
(zeatin) or with [6-benzyladenine] (BA). After 45 d of culturing, clumps of green bulbous structures containing small adventitious
buds (clusters) were generated in all explants cultured with 10 μM zeatin under a photon flux density of 65 μmol m−2 s−1. For subsequent shoot differentiation, clusters were transferred onto medium containing 1 μM zeatin. After 60 d of culturing, 30% of clusters generated from petiole explants developed into plants. The regenerated plantlets
were successfully acclimatized and all survived and grew well. No morphological abnormalities were observed. 相似文献
4.
Summary Direct somatic embryo formation and plantlet regeneration was achieved from immature leaflets of chickpea (Cicer arietinum L.). Optimal somatic embryogenesis was obtained when immature leaflets were exposed to media supplemented with 15 μM 2,4-dichlorophenoxyacetic acid (2,4-D) for 7 d, to 2000 μM 2,4-D for 3 d, and to 50 μM 2,4-D for 10 d, followed by transfer onto Murashige and Skoog (MS) basal medium. Exposure of explants to high 2,4-D levels
(200–2000 μM) for 3 d produced bottle-shaped embryos, while exposure to low 2,4-D levels (<50 μM) and 50–2000 μM for 10 d produced spherical-shaped embryos. Two percent of embryos converted into plants upon culture on MS medium containing
15 μM gibberellic acid and 1 μM 3-indolebutyric acid. All regenerated plants were phenotypically normal. 相似文献
5.
Summary The present study reports that a revised nutrient concentration in the basal medium improved shoot bud induction and subsequent
plant regeneration in barley (Hordeum vulgare L. var. BL-2). Cultures were raised from immature embryos on MSB5 medium supplemented with picloram. Concentrations of five nutrients were varied. The effect of these nutrients was investigated
on (1) induction, (2) induction and subculture, and (3) induction, subculture and regeneration stages. The basal MSB5 medium was not optimal for each phase of barley culture. Decreased ammonium nitrate, increased potassium dihydrogen phosphate,
sodium molybdate, cobalt chloride, and addition of glycine enhanced shoot bud induction and plant regeneration. The different
media that were optimal for immature embryo culture were: MSB5 medium supplemented with 20.70 μM picloram, 10.30 mM NH4NO3, 6.25 mM KH2PO4, 2.06 μM Na2MoO4, 0.55 μM CoCl2, and 26.64 μM glycine (for induction); MSB5 medium supplemented with 12.47 μM picloram, 10.30 mM NH4NO3, and 0.55 μM CoCl2 (for subculture); and MSB5 medium supplemented with 0.2 μM picloram and 10.3 mM NH4NO3 (for regeneration). Primary cultures required 6wk (without transfer) for morphogenic callus formation. Callus required 4wk
of subculture and another 4wk on regeneration medium for optimal plant regeneration. The revised medium could also promote
regeneration of the recalcitrant barley genotype RD-2552. Histological analysis showed that the major pathway of differentiation
was through shoot bud formation. 相似文献
6.
A simple suspension culture system of Platycerium bifurcatum was developed where sporophytes could be regenerated directly from leaf cells or indirectly through an aposporous gametophyte
stage under the same culture conditions. Single cells and aggregates of up to 100 cells developed aposporous gametophytes
which later gave rise to sporophytes. Such gametophytes started apogamy when they were mostly less than 0.7 mm in length,
bearing only rhizoids. In most cases, only one sporophyte was regenerated from one gametophyte. Aggregates of 500–1000 or
more cells, on the other hand, regenerated sporophytes directly. Intercellular interaction was considered to be the physiological
cause, and the separation of leaf cells to a certain degree drove the cells to embark on different regeneration paths. It
is suggested that the possible existence of a threshold size of cell aggregates separates the two regeneration patterns.
Received: 3 March 1997 / Revision received: 11 April 1997 / Accepted: 3 June 1997 相似文献
7.
Summary Explants derived from adventitious buds, rhizomes, stems, and leaves of a medicinal plant, Polygonatum cyrtonema, were studied for plantlet regeneration, and only adventitious bud explants were able to be regenerated into plantlets. Regeneration
was also accompanied by the formation of rhizome-like tissue, the medicinal portion of the plant. The optimum hormone combination
for plantlet regenertion was 4.44 μM benzyladenine plus 2.26 μM 2,4-dichlorophenoxyacetic acid, at which new adventitious buds were obtained from 96.6% of the adventitious bud explants,
with an average of 5.2 buds per explant. The best medium for root induction was half-strength Murashige and Skoog medium with
4.57 μM α-naphthaleneacetic acid, as 92% of regenerated buds rooted. Regenerated plantlets were successfully transferred to a greenhouse
with 86% survival. Histological observation indicated that new adventitious buds originated from the superficial meristematic
cell cluster of the granular callus induced from adventitious bud explants via organogenesis. 相似文献
8.
Filomena Calixto M. Salomé Pais 《In vitro cellular & developmental biology. Plant》1997,33(2):119-124
Summary
Pinus pinaster plants were regenerated from cotyledons excised from in vitro germinated seeds and axenically cultured on induction medium (GMD). 6-Benzyladenine (2.2 μM) induced the highest frequency of direct bud formation from cotyledons. An average of 13.1 ± 2.1 elongated shoots per cotyledon
was obtained. Germination time influenced shoot induction, and the organogenic potential decreased with explant age. Cotyledons
remained for 21 d on induction medium, and in order to promote adventitious shoot elongation, they were transferred to Gupta
and Durzan’s DCR medium without growth regulators, containing 0.5% (wt/vol) activated charcoal and 3% (wt/vol) sucrose. Rooting
was achieved by application of an indole-3-butyric acid, (396.6 μM) pulse (24 h at 4° C), followed by transfer to a sterile mixture of peat plus perlite (1:1 vol/vol). Ninety-eight to 100%
of the regenerated plants were successfully acclimatized. All plants have survived after transfer to the field. 相似文献
9.
《Comptes rendus de l'Académie des sciences. Série III, Sciences de la vie》1999,322(12):1105-1111
A method for a high frequency and direct in vitro bud regeneration of a woody species, the trifoliate orange (Poncirus trifoliata L. Raf), was designed. Transverse thin cell layer (tTCL) explants excised from the stem internodes of 1-year-old young plants of P. trifoliata regenerated bud in vitro on a medium containing 6-benzylaminopurine (BAP 1-50 μM) and N-phenyl-N'-1,2,3-thidiazol-5-ylurea (thidiazuron, TDZ) (0.1–10 μM). The optimal concentrations for bud induction were 25 μM BAP and 1 μM TDZ leading to 87 and 72 % of responsive tTCLs and 24 and 15 buds per tTCL, respectively. A higher percentage of responsive tTCLs and a higher frequency of bud regeneration were obtained with BAP and TDZ combined. With a combination of 10 μM BAP and 1 μM TDZ, 90 % of responsive tTCLs forming 37 buds per tTCL were obtained. Shoot elongation occurred after a transfer onto a medium containing 1 μM GA3. Rooting of individual shoot was induced using 5 μM NAA. One hundred per cent of rooted shoots developed normally after transfer to the greenhouse; no phenotype variation was observed. High numbers of regenerated viable plants can be produced directly without callus formation from tTCL after 9 weeks of culture. 相似文献
10.
The micropropagation of adult Cleistanthus collinus was accomplished. The nodal segments from terminal twigs of a 15-year-old tree and basal sprouts of a comparable chronological
age were used for initiating shoot bud cultures. Washing explants with sterile mixtures of citric acid (520.5 μM) and PVP 40 (3.75 μM) three to four times controlled the leaching of brown inhibitory substances into the establishment medium. Axillary shoots
proliferated best on MS medium containing citric acid (104.1 μM), and PVP 40 (12.5 or 25 μM) supplemented with 0.44 μM BA. The number of new shoots from nodal segments of explants placed on MS medium supplemented with 0.44 μM BA increased when the remaining lengths of nodal segments were transferred to fresh medium after the longer microshoots were
harvested. The microshoots derived from basal sprouts rooted best (50%) when treated with 11.4 mM IAA for 2 min, whereas only 40% of the microshoots derived from terminal twigs produced roots after a 2-min exposure to 28.5
mM IAA. The placement of BA-soaked agar cubes on the apex-decapitated shoots controlled shoot-tip necrosis considerably. In
general, explants from basal sprouts were more suitable than terminal twig explants for the micropropagation of adult trees
of C. collinus.
Received: 26 February 1997 / Revision received: 13 September 1997 / Accepted: 29 September 1997 相似文献
11.
Y. Sahoo S. K. Pattnaik P. K. Chand 《In vitro cellular & developmental biology. Plant》1997,33(4):293-296
Summary An efficient protocol for in vitro propagation of an aromatic and medicinal herb Ocimum basilicum L. (sweet basil) through axillary shoot proliferation from nodal explants, collected from field-grown plants, is described.
High frequency bud break and maximum number of axillary shoot formation was induced in the nodal explants on Murashige and
Skoog (1962) medium (MS) containing N6-benzyladenine (BA). The nodal explants required the presence of BA at a higher concentration (1.0 mg·l−1, 4.4 μM) at the initial stage of bud break; however, further growth and proliferation required transfer to a medium containing BA
at a relatively low concentration (0.25 mg·gl−1, 1.1 μM). Gibberellic (GA3) at 0.4 mg·l−1 (1.2 μM) added to the medium along with BA (1.0 mg·l−1, 4.4 μM) markedly enhanced the frequency of bud break. The shoot clumps that were maintained on the proliferating medium for longer
durations, developed inflorescences and flowered in vitro. The shoots formed in vitro were rooted on half-strength MS supplemented with 1.0 mg·l−1 (5.0 μM) indole-3-butyric acid (IBA). Rooted plantlets were successfully acclimated in vermi-compost inside a growth chamber and
eventually established in soil. All regenerated plants were identical to the donor plants with respect to vegetative and floral
morphology. 相似文献
12.
Summary A protocol for large-scale propagation of Phragmites communis Trin. by adventitious bud formation and plant regeneration was established. Adventitious buds were induced through either
the indirect pathway or the direct pathway from stem explants of Phragmites communis. In the indirect pathway, it was essential to decrease the level of 2,4-dichlorophenoxyacetic acid from 9.1 to 0.5 μM to induce adventitious buds and achieve plant regeneration. In the direct pathway, the effects of different benzylaminopurine
(BA) concentrations in the medium, and different positions of the explants, on adventitious bud formation were determined.
Murashige and Skoog (MS) medium supplemented with 5.4μM α-naphthaleneacetic acid (NAA) and 53.4 μM BA, and the bottom part of stem explants were most responsive for the differentiation of adventitious shoot buds. The highest
differentiation frequency was 20–30 adventitious shoot buds per stem node tissue. Elongation and proliferation of adventitious
buds were achieved on MS medium supplemented with 13.3 μM BA and 5.4 μM NAA. Shoots were rooted in liquid half-strength MS medium with 5.4 μM NAA+4.9 μM indole-3-butyric acid. Rooted plants survived (87.5%) and grew well after transfer into soil for 4 wk. More than 20 000 regenerated
plants of a salt-tolerant variant line of Phragmites communis have been produced. This protocol is useful for clonal micropropagation and possibly for Agrobacterium- mediated gene transfer in P. communis. 相似文献
13.
Y. E. Choi D. C. Yang E. S. Yoon K. T. Choi 《In vitro cellular & developmental biology. Plant》1999,35(3):210-213
Summary Cotyledon explants of Panax ginseng at various developmental stages were cultured on Murashige and Skoog (MS) medium with 0.5 μM indole butyric acid and 8.8 μM N6-benzyladenine. Upon culturing of cotyledon explants from mature zygotic embryos, 34% of the explants formed somatic embryos,
and 46% formed adventitious shoots. In the cotyledon explants from 1-wk-old seedlings, embryo axis-like shoots and roots developed
at a high frequency (79%) near the excised portion of the cotyledon base. The developmental pattern of embryo axis-like organ
formation was structurally different from that of somatic embryos and adventitious shoots but similar to that of parts of
the embryo axis of zygotic embryos. In the early stages of embryo axis-like organ formation, epicotyl-like shoot primordia
were developed directly from the cotyledon base after 2 wk of culture; subsequently roots developed near the base of the epicotyl-like
shoots and eventually regenerated into plantlets with both shoots and roots. The frequency of embryo axis-like organ formation
declined as the growth of seedlings proceeded. In addition, the frequency of somatic embryo and adventitious bud formation
rapidly declined with the age of the cotyledons. Plant regeneration via embryo axis-like organ formation might be a new pattern
of morphogenesis in P. ginseng cotyledon culture. 相似文献
14.
Summary A highly efficient protocol for callus induction and plant regeneration in Sorghum bicolor was developed by varying the concentrations of copper (0.1, 0.3, 0.5, 0.7, 1, 1.5, 2.5 μM) in Murashige and Skoog (MS) medium. The mature embryos of Sorghum bicolor were cultured on MS medium containing 2,4-dichlorophenoxyacetic acid (9μM), kinetin (2.3 μM), and 3% (w/v) sucrose for embryogenic callus induction. Plant regeneration from this callus occurred on MS medium containing
kinetin (9.2 μM) and indole-3-acetic acid (2.85 μM). A much greater response was noted on these media with higher levels of copper. Frequency of plant regeneration and number
of regenerants dramatically increased with an optimal amount of copper (2 μM) in the MS medium. Rooting of the regenerated shoots readily occurred on half-strength MS medium supplemented with α-naphthaleneacetic
acid (10.7 μM) and 3% (w/v) sucrose. Well-developed plantlets were transferred to the field where 100% survival and normal seed setting
was noted. 相似文献
15.
Wennuan Liu C. Ernie Chilcott Richard C. Reich Gary M. Hellmann 《In vitro cellular & developmental biology. Plant》2000,36(3):201-206
Summary The present work provides a system for regeneration of clary sage, (Salvia sclarea L.) via organogenesis using plant tissue culture techniques in a multistage culturing medium containing 2,4-dichlorophenoxyacetic
acid (2,4-D) (9.05–181.00 μM). A higher frequency of organogenic tissue initiation was obtained from immature zygotic embryo cotyledons (IZEC) 2–3 wk
after pollination on the medium supplemented with 9.05 μM 2,4-D. The organogenic tissues were then proliferated on media containing both indole-3-acetic acid (IAA) and 6-benzylaminopurine
(BA). Organogenic lines were established via selection, isolation and continuous subculture of organogenic tissues on a medium
containing 22.19 μM BA and 2.85 μM IAA. Shoots were regenerated from both the proliferated tissues and IZEC, and propagated in the presence of IAA or α naphthaleneacetic
acid (NAA), BA and gibberellic acid (GA3). Although roots were induced from regenerated shoots on the media containing a low concentration of IAA, IBA (0.98 μM) in combination with desiccation of regenerated shoots with a stem ∼10 mm in length promoted more and stronger root formation.
After the root system was well established (20 mm in length), the regenerated plants were transferred to soil in plastic pots
for further growth and production of R1 seeds in the greenhouse. 相似文献
16.
Luis Pedro Barrueto Cid Rolf Dieter Illg Aquiles E. Piedrabuena 《In vitro cellular & developmental biology. Plant》1994,30(3):150-155
Summary Aiming at the genetic improvement of garlic cultivars, a cell suspension protocol was established which includes the induction
of friable callus, establishment of cells in liquid medium, plating, regeneration, and bulb formation. Calluses of various
textures from compact to friable and from green to yellowish were obtained by culturing explants excised from inner leaves
of garlic bulbs on Marashig-Shoog (MS) medium with 2,4 dichlorophenoxy acetic acid (2,4-D), (1.1 mg/liter [5.0 μM]), picloram
(1.2 mg/liter [5.0 μM]), and kinetin (2.1 mg/liter [10 μM]). Friable callus occurred on MS-A contained 2,4-D alone (1.0 mg/liter
[4.52 μM]) and this callus was used to develop cell suspension cultures, which were maintained in liquid MS-B medium with a 2,4-D/benzyl
adenine (BA) (0.5 mg/liter [2.25 μM]: 0.5 mg/liter [2.22 μM]) ratio. High plating efficiency was obtained on MS-C medium with different naphthalene acetic acid/BA combinations. Regeneration
occurred after transfer of the caulogenic mass to MS-C medium containing 10 mg/liter (74.02 μM) and 20 mg/liter (148.04 μM) adenine for 60 days, followed by transfer to adenine-free medium. Plantlets transplanted to soil showed normal phenology.
Shoots grown on modified MS medium supplemented with indolylbutryic acid (3.0 mg/liter [14.7 μM]) stimulated bulb formation by 30 days in culture. 相似文献
17.
A rapid and reliable micropropagation method was established for Spathoglottis plicata. Nodal and leaf explants dissected from 8-month-old pot-grown seedlings were cultured on charcoal-amended Murashige and Skoog
medium supplemented with 16 combinations of α-naphthaleneacetic acid (NAA) and 6-benzylaminopurine (BA) at concentrations
of 0.54–10.74 μm. Regeneration of protocorm-like bodies (PLBs) and subsequent plantlet development were observed from 98.5% of the nodal explants.
Only 6.5% of leaf explants and occasionally some root segments (dissected from regenerated plantlets) were able to produce
PLBs and then plantlets. The optimum plant growth regulator (PGR) combination for maximal PLB regeneration was 5.37 μm NAA and 0.44 μm BA. The best combination of PGR for plantlet development was 2.69–10.74 μm NAA and 8.88 μm BA. The NAA to BA ratios for maximal PLB induction and plantlet development were 12.2 and 0.3–1.2, respectively. Regenerated
PLBs and plantlets, when cut into pieces of less than 1 mm and subcultured onto the above media, regenerated new PLBs and
plantlets in another 3 months.
Received: 20 February 1997 / Revision received: 27 May 1997 / Accepted: 16 June 1997 相似文献
18.
Shoot bud regeneration from Petunia leaf disks was inhibited when they were cultured with the demethylating agents, 5-azacytidine (AzaC) and 5-aza-2′-deoxycytidine
(AzadC), in shoot induction (SI) medium. Explants induced shoot primordia if they were transferred after 1 week from the medium
containing the drugs to medium without drugs. The fresh weight of leaf disks cultured on SI medium for 2 weeks in the presence
of the drugs was 60–80% lower when compared to control shoot-forming cultures. Internode length was reduced when shoots were
transferred to phytohormone-free Murashige and Skoog medium containing the drugs. However, no other morphological abnormalities
were seen in these shoots, even at 20 μm AzaC or 5 μm AzadC. Coupled restriction enzyme digestion (with HpaII and MspI) and random amplification of genomic DNA was performed to detect the level of methylation of CCGG sites in the DNA of the
explants exposed to AzaC and AzadC. Over 15 amplified bands were detectable in the control. Five of these bands were absent
in the amplified products when digested DNA from the drug-treated explants was used as the template, showing that hypomethylation
of DNA had occurred. This suggests that inhibition of shoot bud formation in the presence of the drugs AzaC and AzadC may
be due to the altered methylation status.
Received: 7 January 1997 / Revision received: 17 February 1997 / Accepted: 1 March 1997 相似文献
19.
Adventitious bud formation from the vegetative buds of the flower stalks of Phalaenopsis occurred on Vacin and Went medium with 15% coconut water and 5 to 40 μM thidiazuron (TDZ) or 40 μM N6-benzylaminopurine. The highest efficiency of induction was achieved with 5 or 10 μM TDZ. Adventitious buds developed into shoots on VWC medium. TDZ was more effective than BAP in stimulating the axillary buds of intact shoots to develop. Regenerated shoots rooted after about two months of culture on VWC medium with 1% sucrose. Shoot tips excised from the regenerated shoots initiated protocorm-like bodies after two months of culture on VWC medium. 相似文献
20.
A. E. Jofre-Garfias N. Villegas-Sepúlveda J. L. Cabrera-Ponce R. M. Adame-Alvarez L. Herrera-Estrella J. Simpson 《Plant cell reports》1997,16(12):847-852
Mature embryos of Amaranthus hypochondriacus (amaranth) were used to develop an in vitro culture system for plant regeneration and genetic transformation. Plants were regenerated from embryo-derived callus cultivated
on Murashige and Skoog medium supplemented with 10 μM 2,4-dichlorophenoxyacetic acid or 3,6-dichloro-2-methoxybenzoic acid and 10% coconut liquid endosperm. Transgenic plants
were obtained by inoculation of mature embryo explants with a disarmed Agrobacterium strain containing the plasmid pGV2260(pEsc4), which carried the genes encoding neomycin phosphotransferase type II and β-glucuronidase. The presence of transgenes in the genome of transformed amaranth plants and their progeny was demonstrated
by Southern blot hybridization. Tissue specific and light-inducible expression directed by a pea chlorophyll a/b-binding protein
promoter was observed in transgenic amaranth plants and their progeny.
Received: 30 December 1996 / Revision received: 14 May 1997 / Accepted: 3 June 1997 相似文献