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1.
Using cotyledon explants excised from seedlings germinated in vitro, an efficient plant regeneration system via organogenesis was established for bottle gourd (Lagenaria siceraria Standl.). Maximum shoot regeneration was obtained when the proximal parts of cotyledons from 4-day-old seedlings were cultured on MS medium with 3 mg/l BA and 0.5 mg/l AgNO3 under a 16-h photoperiod. After 3–4 weeks of culture, 21.9–80.7% of explants from the five cultivars regenerated shoots. Adventitious shoots were successfully rooted on a half-strength MS medium with 0.1 mg/l IAA for 2–3 weeks. Flow cytometric analysis revealed that most of the regenerated plants derived from culture on medium with AgNO3 were diploid. 相似文献
2.
Conditions of transformation and regeneration of `Induka' and `Elista' strawberry plants 总被引:3,自引:0,他引:3
Gruchała Agnieszka Korbin Małgorzata Żurawicz Edward 《Plant Cell, Tissue and Organ Culture》2004,79(2):153-160
Efficiency of plants' transformation depends on many factors. The genotype, applied techniques and conditions of plant's modification and modified plant regeneration are the most important among them. In our studies regeneration and transformation conditions for two strawberry cultivars were determined and compared. Plants were transformed by Agrobacterium tumefaciens LBA4404 strain containing plasmid pBIN19 with nptII and gus-reporter genes. Experiment was carried out on more than 1300 leaf explants from each cultivar. Generally, `Induka' plants characterized with higher regeneration potential than `Elista'. The highest number of regenerated shoots was obtained on MS medium with 0.4 mg l –1 IBA and 1.8 mg l–1 BA (3.5 and 1.8 shoots/explant for `Induka' and `Elista', respectively). After plant transformation number of regenerated, transgenic shoots was higher for `Elista' (on the average: 8.3 shoots/100 explants). The number of transgenic `Induka' shoots, obtained at the same conditions, was twice lower (4.2). Simultaneously `Induka' plants needed higher kanamycin concentration for transgenic explants selection than `Elista' (25 mg l–1). Preliminary incubation of A. tumefaciens in LB or MS medium with acetosyringone and IAA resulted in increasing transgenic shoots number (per 100 explants: `Induka' 4.5, `Elista' 8.0–9.5 shoots). After using untreated bacteria for plants' transformation, number of transgenic plants varied (dependently on cultivar) from 3.8 to 7.0/100 explants. Applying LB or MS as basic medium as well as adding tobacco plant extract to these media did not significantly influence transformation efficiency. 相似文献
3.
We have developed a new Agrobacterium-mediated transformation method for the low-frequency-regenerating pear (Pyrus communis L.) cvs. Silver bell and La France. Leaf sections derived from in vitro shoots were initially used for the transformation procedure. Under optimum transformation conditions, which included culture and selection on 30 mg/l kanamycin (Km) combined with 500 mg/l sulbenicillin, a 3.2% transformation efficiency was obtained for cv. Silver bell, but no transformants of La France were obtained because of the very low regeneration frequency. Axillary shoot meristems were then examined as potential explants for La France. Selection in 5 mg/l Km and 375 mg/l carbenicillin resulted in transformed shoots being produced at an efficiency of 4.8%, and the apparent white Km-sensitive shoots were not formed during a 2-year subculture on micropropagation medium containing 50 mg/l Km. Therefore, transformations using axillary shoot meristems may be an alternative method for pear cultivars recalcitrant to regeneration from leaf sections. 相似文献
4.
Several culture conditions were examined for promoting efficient plant regeneration from explants of Gentiana. Adventitious shoot regeneration from leaf explants of cv. WSP-3 was very superior on MS medium, compared to B5 medium, supplemented with four cytokinins (TDZ, 4PU-30, BA and zeatin). An auxin / cytokinin combination was required for regeneration. TDZ was the most effective cytokinin, while NAA was more effective than IAA or 2,4-D. Optimum conditions for regeneration from explants (leaf, stem and root) of cv. WSP-3, evaluated in terms of regeneration frequency and number of regenerated shoots per explant, were TDZ and NAA in combination, 5–10 mg/l and 0.1 mg/l for leaf and stem explants, and 10 mg/l and 1 mg/l for root explants, respectively. Application of these conditions to eight other commercial cultivars resulted in 30–100% regeneration from leaf explants. The number of chromosomes in each of ten regenerated plants of each cultivar was diploid, 2n=26. Shoots regenerated in vitro were rooted in phytohormone-free medium and transferred to soil.Abbreviations MS medium
Murashige and Skoog's medium (Murashige and Skoog 1962)
- B5 medium
Gamborg B5 medium (Gamborg et al. 1968)
- BA
6-benzylaminopurine
- TDZ
N-phenyl-N'-1,2,3-thiadiazol-5-yl urea
- 4PU-30
N-(2-chloro-4-pyridyl)-N'-phenylurea
- 2,4-D
2,4-dichlorophenoxyacetic acid
- IAA
indole-3-acetic acid
- NAA
1-naphthaleneacetic acid 相似文献
5.
F. Scaramuzzi G. Apollonio S. D’Emerico 《In vitro cellular & developmental biology. Plant》1999,35(3):217-221
Summary Leaf explants of Sinningia speciosa were cultured in vitro on Murashige and Skoog (MS) basal medium with various growth substances in order to regenerate shoots.
On MS medium supplemented with indoleacetic acid (IAA) and kinetin, 80% of the explants produced green callus and 25 to 30
shoots with roots per explant. On MS supplemented with IAA and N6 benzyladenine (BA), 80% of the explants produced green callus and 40 to 50 shoots per explant but lacked roots. After 3–4
mo., these shoots were removed from the initial explants and transferred separately onto MS supplemented with indolebutyric
acid for their elongation and successive rooting (3 mo.). Histological studies showed that the callus was associated with
mesophyll cell layers, primarily with the spongy parenchyma. The shoots regenerated at the callus surface and were associated
with newly differentiated vascular areas. Recurrent regenerations were obtained from leaf explants or apical meristems excised
from shoots of the previous subcultures. These explants, as compared to initial cultures, had a high frequency of regeneration
and also produced more shoots per explant. Chromosome numbers of root tip cells of the mother plant and of all in vitro-regenerated
plants remained constant: 2n=26. 相似文献
6.
Pradeep K. Agarwal Rajinder S. Ranu 《In vitro cellular & developmental biology. Plant》2000,36(5):392-397
Summary The in vitro plant regeneration potential of vegetatively propagated geraniums (Pelargonium x hortorum) has been investigated. Using various combinations of growth regulators and a choice of different explants, a regeneration
protocol has been developed to raise in vitro plantlets from young petiole and leaf explants from three different cultivars of geraniums. In all three cultivars, very
young petiole explants exhibited a higher regeneration potential as compared with leaf explants. Regeneration efficiencies
were found to be highly dependent on the cultivar, with cv. Samba showing the highest regeneration potential, followed by
cvs. Yours Truly and then Sincerity. Samba also showed the highest number of shoots from both the petiole [57 shoot buds per
petiole explant in the presence of 3 μM zeatin and 1 μM indole-3-acetic acid (IAA) and leaf explants (43 shoots per leaf explant with 10 μM zeatin and 2 μM IAA). Shoot buds transferred to Murashige and Skoog (MS) medium supplemented with 0.44 μM N6-benzyladenine and 0.11 μM IAA grew vigorously and attained 1–2 cm in length in 3–4 wk. These shoots rooted with 100% efficiency on MS basal medium,
and plants developed that showed normal growth and flowering under greenhouse conditions. 相似文献
7.
Esmaeil Ebrahimie A. A. Habashy M. Mohammadie-Dehcheshmeh M. R. Ghannadha B. Ghareyazie B. Yazdi-Amadi 《In vitro cellular & developmental biology. Plant》2006,42(5):455-460
Summary A rapid and one-step protocol for direct regeneration of shoots from cumin embryo explants has been developed. Embryo explants
with shoot meristems were cultured on shoot regeneration medium for 15–22 d. After embryo culture, shoots were regenerated
from the area adjacent to the region between the cotyledons and embryo axis within 2 wk, without any intermediate callus phase.
Shoot proliferation and elongation were achieved on shoot regeneration medium without subculture. Among the different combinations
of 6-benzylaminopurine, α-naphthaleneacetic acid (NAA), and indole-3-acetic acid (IAA) tested, 0.8 mgl−1 (4.3 μM) NAA in combination with 0.3 mgl−1 (1.71 μM) IAA in the B5 medium resulted in the most efficient direct shoot regeneration. No significant difference was detected for
the number of regenerated explants when different heterogeneous endemic varieties were compared. This plant regeneration procedure
was applicable to different cumin genotypes and regenerated plants were phenotypically normal. 相似文献
8.
Jayanand B. Sudarsanam G. Sharma Kiran K. 《In vitro cellular & developmental biology. Plant》2003,39(2):171-179
Summary An efficient and reproducible protocol for the regeneration of shoots at high frequency was developed by using explants derived
from the axillary meristems from the cotyledonary nodes of in vitro-germinated seedlings of chickpea (Cicer arietinum L.). Culture conditions for various stages of adventitious shoot regeneration including the induction, elongation, and rooting
of the elongated shoots were optimized. The medium for synchronous induction of multiple shoot buds consisted of Murashige
and Skoog basal medium (MS) with low concentrations of thidiazuron (TDZ), 2-isopentenyladenine (2-iP), and kinetin. Exclusion
of TDZ and lowering the concentration of 2-iP and kinetin in the elongation medium resulted in faster and enhanced frequency
of elongated shoots. Cultivation of the stunted shoots on MS with giberellic acid (GA3) increased the number of elongated shoots from the responding explants. pH of the medium played a very crucial role in the
regeneration of multiple shoot buds from the explants derived from cotyledonary nodes. A novel rooting system was developed
by placing the elongated shoot on a filter paper bridge immersed in liquid rooting medium that resulted in rooting frequency
of up to 90%. A comprehensive protocol for successful transplantation of the in vitro-produced plants is reported. This method will be very useful for the genetic manipulation of chickpea for its agronomic improvement. 相似文献
9.
Summary We have developed efficient methods for plant regeneration, via both embryogenesis and organogenesis, of Smooth Cayenne pineapple,
Ananas comosus (L.) Merr. Leaf bases and core (stem) sections of in vitro shoots, produced from culture of crown tip meristem, were used as explants for plant regeneration as follows: (1) Leaf base
and core section explants cultured on Murashige and Skoog (MS) medium containing 41 μM 4-amino-3,5,6-trichloropicolinic acid (picloram, P) or thidiazuron (T)/P combinations produced embryogenic tissues. Different
types of embryogenic tissues (friable emryogenic tissue, embryogenic cell cluster, and chunky embryogenic tissue) have been
developed with varying properties in terms of growth rate and state of development. The embryogenic tissues regenerated shoots
upon culture on MS medium containing 13 μM 6-benzylaminopurine (BA) and 1μM α-naphthaleneacetic acid (NAA) followed by culture on MS medium containing 4 μM BA. (2) Crown tip meristems cultured on MS medium containing 13 μM BA followed by leaf explants cultured on MS medium with 27 μM NAA and 1 μM BA produced shoots via direct organogenesis. (3) Explants cultured on MS medium containing 5 μM T and 0.5 μM indole-3-butyric acid (IBA) produced nodular globular structures, which produced shoots upon culture on MS medium containing
1 μM BA and 1 μM gibberellic acid. Shoots obtained from all of the above methods were rooted in half-strength MS medium containing 3 μM NAA and 2.5 μM IBA. Plants were transferred to the greenhouse or shipped to Costa Rica for field trials. Somatic embryo-derived plants exhibited
21 % spininess, and organogenic-derived plants exhibited 5% spininess in the field trials. 相似文献
10.
Summary A facile procedure has been developed to regenerate white clover (Trifolium repens L.) plants, rapidly and directly from cotyledon explants of 3 day old seedlings. Scanning electron microscopy and histological sectioning demonstrated that shoot meristems developed from individual epidermal cells on the adaxial surface of the cotyledonary stalk, proximal to the site of excision. Initial cell divisions occurred after 2 days of culture and regenerated plants were transferred to soil within 6–8 weeks. Regenerated plants were normal, flowered and set seed. The highest shoot regeneration frequency (an average of 20 shoots per cotyledon) was obtained using an MS based medium containing 1.0 mg 1-1 6-benzylaminopurine and 0.05 mg 1-1 -napthaleneacetic acid. A similar regeneration frequency was obtained from cotyledon explants taken from eight different white clover cultivars.Abbreviations BAP
6-benzylaminopurine
- NAA
-napthaleneacetic acid
- MS
Murashige and Skoog medium 相似文献
11.
Genotypic response of cowpea Vigna unguiculata (L.) to in vitro regeneration from cotyledon explants
M. S. Brar J. M. Al-Khayri T. E. Morelock E. J. Anderson 《In vitro cellular & developmental biology. Plant》1999,35(1):8-12
Summary A plant regeneration system applicable to 17 cowpea genotypes was developed. Cotyledons were initiated on 1/3 MS medium containing
15 to 35 mg N6-benzyladenine (BA) per 1 (66.6 to 155.3 μM) for 5 to 15 d. For shoot regeneration, the explants were transferred to a medium containing 1 mg BA per 1 (4.4 μM). Within 1 wk, shoot formation was visible at the proximal end of the cotyledons. Regeneration percentages (1% to 11%) and
the numbers of shoots (4 to 12 per explant) were significantly influenced by genotype. Culture duration and BA concentration
in the initiation stage significantly affected regeneration capacity. Explants initiated on media containing 15 mg BA per
1 for 5 d resulted in the highest percentage of explants capable of regeneration. Conversely, the highest number of shoots
was obtained from explants initiated on media supplemented with 35 mg BA per 1. Whole plants were obtained on a plant growth
regulator-free medium. To our knowledge, this is the first report of plant regeneration from U.S. commercial cowpea cultivars
and breeding lines. This system is adaptable to diverse cowpea genotypes and will facilitate cowpea genetic transformation.
Published with the approval of the Director of the Arkansas Agricultural Experiment Station. 相似文献
12.
C. Pugliesi M. Rabaglio F. Cecconi S. Baroncelli 《Plant Cell, Tissue and Organ Culture》1992,28(2):125-128
Different plant explants of Persian buttercup (Ranunculus asiaticus L.) were screened for callus induction and adventitious shoot regeneration on different media to establish totipotent cultures. Murashige & Skoog (MS) medium was used, supplemented with different concentrations of the following growth regulators: kinetin, benzyladenine (BA), naphthaleneacetic acid (NAA) and indoleacetic acid (IAA). Callus was induced and adventitious buds regenerated only from cotyledonary explants after 4–5 weeks. Subculture of the regenerated buds on the same basal medium in presence of gibberellic acid (GA3) and BA produced well-organized shoots. Rooting was obtained by transferring shoots to growth regulator-free MS medium. A high rate of shoot multiplication has been achieved on medium with high concentration of kinetin and long-day photoperiod. Finally the plants were successfully transferred to soil and grown in a greenhouse. 相似文献
13.
Methods are described for obtaining explants which produce adventitious shoots, for subsequent stimulation of rooting and then transplanting using six commercial sugar-beet cultivars. The rate of adventitious shoot regeneration from petioles or intact leaf explants was affected by the source of donor plants, cytokinin type (BAP or Kin) and concentration and cultivar. Increasing the sucrose concentration of the medium from 3% to 5% or 8% had no apparent effect. Adventitious shoots could be produced directly from callus formed on the base of the petioles. In general adventitious shoots were produced on either the concave surface of the petiole or from the callus, occasionally simultaneously on both, and on the convex surface of the petiole in intact leaf explants. The highest rooting rate with 3% sucrose and 1.0 mg l–1 NAA was obtained using half-strength MS medium. There was considerable variation in the propagules from petioles or callus indicating that this system may provide valuable somaclonal variation.Abbreviations BAP
benzylaminopurine
- IBA
indole-3-butyric acid
- GA3
gibberellic acid
- MS
Murashige and Skoog medium
- NAA
naphthaleneacetic acid
Author for correspondence 相似文献
14.
An effective protocol has been developed for plant regeneration from cotyledon explants of Swainsona salsula Taubert (Saline swainsona), a medicinal and agronomic shrub. Adventitious shoots were obtained from 83.2% of cotyledon explants
from 3-day seedlings cultured on Murashige and Skoog (MS) medium containing 2.0 mg l−1 thidiazuron (TDZ), with an average of 9.3 shoots per explant. Individual elongated shoots were rooted on half strength MS
medium supplemented with 2.0 mg l−1 indole-3-butyric acid (IBA), with 59.3% success. Regenerated plants with well developed shoots and roots were successfully
transferred to soil, without detectable variants. Histological observation revealed that shoots developed from cotyledon explants
via organogenesis, with little callus.
This revised version was published online in June 2006 with corrections to the Cover Date. 相似文献
15.
Two plant regeneration methods applicable to Leucaenaleucocephala were developed. In the first method, involvingorganogenesis via callus formation, cotyledon, hypocotyl and root segments wereinitiated on MS medium containing different concentrations ofN6-benzyladenine (BA), 2,4-dichlorophenoxyacetic acid (2,4-D), andnaphthaleneacetic acid (NAA). Both compact (type I) and friable (type II) calliwere obtained from the cotyledon and hypocotyl explants treated with differentconcentrations of the growth regulators. Shoots were generated only from thefriable calli formed from the cotyledon explants. The calli formed from thehypocotyl explants did not generate shoots and the root explants died withoutforming callus. Cotyledon explants from 3–4 day old seedlings showedmaximum callus induction compared to those from older seedlings. In a secondmethod involving direct organogenesis, excised cotyledons were cultured on 1/2MS medium containing 10–35 mg l–1N6-benzyladenine (BA) for 7–14 days. Transfer of thecotyledonsto regeneration medium containing low BA resulted in callus formation andsubsequent shoot regeneration from the base of the excised cotyledon explants,with up to 100% frequency. Regenerated shoots rooted best on a basal mediumcontaining no growth regulators. 相似文献
16.
Thidiazuron Induced Multiple Shoot Induction and Plant Regeneration from Cotyledonary Explants of Mulberry 总被引:4,自引:0,他引:4
T. Dennis Thomas 《Biologia Plantarum》2003,46(4):529-533
A rapid micropropagation protocol through induced multiple shoots from the cotyledonary explant of mulberry (Morus alba L) is described. The highest number of shoots (20.3) was obtained when explants from 14-d-old embryos were cultured on Murashige
and Skoog (MS) medium supplemented with 7 μM thidiazuron for 45 d. Of the three cultivars used, cv. S-36 was the best followed
by cv. K-2 and S-1. The shoots were transferred to MS medium supplemented with 5 μM 6-benzylaminopurine for elongation. The
elongated shoots were rooted on half strength MS medium containing 1 – 7 μM indole 3-butyric acid or 1-naphthalene acetic
acid. The rooted plants were transplanted to soil with 90 % success. The emerged shoot primordia probably initiated from the
pre-existing meristems since the shoot bud show definite vascular connection to the major vascular tissue.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
17.
This report deals with micropropagation of the critically endangered and endemic Turkish shrub, Thermopsis turcica using callus, root and cotyledonary explants. Callus cultures were initiated from root and cotyledon explants on MS medium
supplemented with 0.5–20 μM NAA or 2,4-D. The root explants were found to be better in terms of quick responding and callusing
percentages as compared to the cotyledons. Organogenic callus production with adventitious roots and shoots were obtained
on MS medium with only NAA. The calli obtained with NAA, root and cotyledonary explants were cultured with BA and kinetin
(2–8 μM) alone or in combination with a low level (0.5 μM) of 2,4-D or NAA. The best regeneration of shoots from root explants
was observed on hormone-free MS medium. NAA with BA or kinetin in the medium improved shoot induction from the calli obtained
with NAA. Maximum percentage of shoots (93.3%), maximum number of shoots (6.2) and maximun length of shoots (8.22 cm) were
achieved from cotyledonary explants at 4 μM BA and 0.5 μM NAA. The presence of 0.5 μM or higher levels of 2,4-D in shoot induction
medium inhibited the regeneration in T. turcica explants. 83% of in vitro rooting was attained on pulsed-IBA treated shoots. The regenerated plants with well developed shoots and roots were successfully
acclimatized. Application of this study’s results has the potential to conserve T. turcica from extinction. 相似文献
18.
Adult plants are known for recalcitrance when it comes to adventitious organ formation and regeneration. Methods used for
regeneration in explants from seedlings of Campanula carpatica failed to work for explants from adult plants of the same species. The present investigation generated efficient regeneration
methods for mature specimens of four species of Campanula, C. carpatica, C. haylodgensis, C. portenschlagiana and C. poscharskyana. Petiole explants from dark-grown in vitro shoot cultures grown from nodal cuttings of adult plants regenerated successfully
(95%), while explants from light-grown in vitro shoot cultures and greenhouse-grown plants regenerated at 12% and zero percentage,
respectively. Dark-treatment, along with media manipulation with plant growth regulators, further enhanced regenerative capacity
of the explants. A MS-based medium containing 10mg l −1 TDZ and 0.25 mg l−1 NAA was the most efficient regeneration medium. Transgenic shoots from C. carpatica (3%) and C. haylodgensis (1%) and transgenic callus from all species were produced using Agrobacterium tumefaciens, and transformation was confirmed by histochemical and Southern blot analyses. Protocols developed in this study may be useful
for achieving efficient regeneration and transformation of recalcitrant adult plants. 相似文献
19.
In this work we report a new method forin vitro chili pepper (Capsicum annuum L.) plant regeneration based on shoot formation from wounded hypocotyls. Chili pepper seeds were surface sterilized and germinated on agar (0.8%) at 25 ± 2°C in the dark. Five factors that may influence shoot regeneration were studied: age of seedlings, hypocotyl wounding site, time elapsed between wounding the hypocotyls and decapitation of seedlings, culture media and cultivars. In order to study the influence of the first three factors on shoot regeneration, the apical, middle or basal hypocotyl regions of seedlings of cv. Mulato Bajio at different stages of development (9, 15, 16, 21 and 28 d old) were wounded with a syringe needle, and the seedlings were cultured on MS semisolid medium without growth regulators at 25 ± 2°C under a 16/8 h light/dark photoperiod (daylight fluorescent lamps; 35 mol m-2 s--1) until decapitation. The seedlings were decapitated (3 mm below the cotyledons) at different times after wounding (0, 2, 4, 10, 12 and 14 d), and each explant was evaluated for bud and shoot formation ( 5 mm in length) at the wounded site after 30 d of incubation. In general, seedlings at the stage of curved hypocotyl (9 d old) wounded in the apical region of hypocotyl were the best explants for shoot regeneration when inoculated on culture medium without growth regulators. Decapitation after wounding also influenced the shoot regeneration efficiency, with 10–14 d being the best period. Up to 90% shoot regeneration in cv. Mulato Bajio was obtained under these conditions. Statistically significant differences were observed for shoot formation among 21 cultivars tested. Regeneration of whole plants was achieved by rooting the shoots with indole-3-butyric acid pulses of 60 mg L–1 for 3 h and then subculturing on MS medium without growth regulators. 相似文献
20.
An efficient micropropagation system for Erigeron breviscapus (vant.) Hand. Mazz., an important medicinal plant for heart disease, has been developed. Shoot organogenesis occurred from
E. breviscapus leaf explants inoculated on a medium supplemented with a combination of plant growth regulators. On average, 17 shoots per
leaf explant were produced after 30 days when they were cultured on MS basal salts and vitamin medium containing 5 μM 6-benzylaminopurine
(BAP) and 5 μM 1-naphthaleneacetic acid (NAA). All the regenerated shoots formed complete plantlets on a medium containing
2.5–10 μM indole-3-butyric acid (IBA) within 30 days, and 80.2% of the regenerated plantlets survived and grew vigorously
in field conditions. Based on the variation in common peaks and the produced amount of the most important bioactive component,
scutellarin, a high performance liquid chromatography (HPLC) fingerprinting system was developed for quality control of these
micropropagated plants. Chemical constituents in E. breviscapus micropropagated plants varied during plant development from regeneration to maturation, the latter of which showed the most
similar phytochemical profile in comparison with mother plants. The regeneration protocol and HPLC fingerprint analysis developed
here provided a new approach to quality control of micropropagated plants producing secondary metabolites with significant
implications for germplasm conservation. 相似文献