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1.
2.
A reproducible system for somatic embryogenesis and plantlet formation of
sandalwood has been developed. A high frequency (100%) of somatic embryos
were induced directly from various explants in MS (Murashige and Skoog,
1962) medium with thidiazuron (1 or 2 M) or
indirectly in medium containing 2,4-D plus thidiazuron. Within 8 weeks,
white globular somatic embryos or friable embryogenic tissue developed on
cultured explants. In S. album the globular somatic
embryos were transferred to MS medium supplemented with IAA (6 M) and kinetin (1 and M) where they
developed further, multiplied and maintained friable embryogenic tissue.
After 15-30 d, mature somatic embryos (1-2 mm) with well-developed
cotyledons were separated and subcultured on to medium containing GA3 (6
M) for germination. Once germinated, elongated somatic embryos
(10-20 mm long) grew further in MS supplemented with lower GA3 (3
M). In S. spicatum, the addition of casein
hydrolysate and coconut milk was necessary for plantlet development from
somatic embryos. From histological studies, it appeared that primary
somatic embryos arose from single cells or had a multicellular origin from
the epidermis or cortical parenchyma. Secondary somatic embryos and friable
embryogenic tissue differentiated from groups of proembryogenic cells from
a superficial layer of the primary somatic embryos.Keywords:
Santalum album, Santalum spicatum, somatic
embryogenesis, histological studies.
相似文献
3.
The stable transformation of embryogenic tissues of Pinus nigra Arn., cell line E104, has been achieved using a biolistic approach. The introduced DNA consisted of the uidA reporter gene under the control of the double CaMV 35S promoter and the nptII selection gene controlled by the single CaMV 35S promoter. Three days after bombardment, putative transformed tissues were selected for continued proliferation on a medium containing 20 mg geneticin l−1. Resistant embryogenic tissue recovery required 10–12 weeks. The integration of the nptII and uidA genes was confirmed by both histochemical/fluorimetric GUS assays and PCR amplification of the inserts in the five geneticin resistant sub-lines of line E104. The activity of the uidA reporter gene in transgenic, embryogenic tissue lines was stable and could be detected after one year of culture. Somatic embryo maturation was, however, poor and no plantlet regeneration could be obtained. 相似文献
4.
Effects of 4 potentially selective agents for transformed cells, 3 antibiotics [kanamycin, geneticin (G418) and hygromycin]
and bialaphos, as well as 2 antibiotics for eliminating Agrobacterium, carbenicillin and cefotaxime on growth and somatic embryogenesis of embryogenic calli of Muscari armeniacum cv. Blue Pearl were evaluated. Callus growth was completely inhibited by 75 mg dm−3 hygromycin or 4 mg dm−3 bialaphos, and somatic embryos were never produced on media containing 25 mg dm−3 hygromycin or 3 mg dm−3 bialaphos. Kanamycin and G418 less inhibited growth and somatic embryogenesis of the calli. On the contrary, carbenicillin
and cefotaxime promoted both callus growth and somatic embryogenesis at all concentrations tested.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
5.
When both cotyledons of light-grown sunflower seedlings (Helianthusannuus L.) were darkened by covering them with aluminium foil,the resulting increase in the rate of elongation of the hypocotylwas closely proportional to the associated reduction in seedlingtranspiration and to the increase in xylem water potential (PX.Covering only one cotyledon induced curvature away from thatside. This response was associated with a higher PX in the vascularbundles connected directly with the covered cotyledon than inthose connected with the illuminated cotyledon. The water contentof peripheral tissues of the hypocotyl below the covered cotyledonwas also higher than that of similar samples from below theilluminated cotyledon. These results are consistent with thehypothesis that the effect of darkening the cotyledons on thegrowth and curvature of the hypocotyl is mediated by hydraulicsignalling, characterized by transmission to the hypocotyl ofan increase in PX resulting from the reduction in cotyledontranspiration. Key words: Cotyledons, Helianthus annuus, hydraulic signalling, hypocotyl, transpiration, water potential 相似文献
6.
Heleen M. van der Maas Eliza R. de Jong Saskia Rueb Lambert A. M. Hensgens Frans A. Krens 《Plant molecular biology》1994,24(2):401-405
Stable transformation of perennial ryegrass (Lolium perenne L.) was achieved by biolistic bombardment of a non embryogenic cell suspension culture, using the hpt and gusA gene. The transformation yielded on the average 5 callus lines per bombardment (1.4×106 cells). Stable integration of the genes into the plant genome was demonstrated by Southern analysis of DNA, isolated from hygromycin-resistant callus lines. The gusA reporter gene, which was regulated by the constitutive promoter of the rice gene GOS2, was expressed in both transient and stable transformation assays, indicating that this promoter is suitable for expression of a transferred gene in perennial ryegrass. Long-term GUS expression was observed in ca. 40% of the callus lines, whereas the other callus lines showed instability after 6 months and 1 year of culture. 相似文献
7.
Short communication. A steep Ca2[IMAGE]-dependence of a K[IMAGE] channel in a unicellular green alga
Bauer C; Plieth C; Hansen U; Simonis W; Schonknecht G 《Journal of experimental botany》1998,49(327):1761-1765
An increase of cytosolic Ca2 in the
unicellular green alga Eremosphaera viridis activities
Ca2-dependent
K channels causing a hyperpolarization of
the plasma membrane. Data from parallel calcium, and potential measurements
were combined with I/V relationships. This yielded a
steep Ca2-dependence of
K channels with a co-operativity of 4 and
an affinity of 300 nM.Key words: Eremosphaera viridis,
plasma membrane, Ca2-dependent
K channel, co-operative binding.
相似文献
8.
A method for Agrobacterium tumefaciens-mediated transformation of Pinus radiata cotyledon explants was developed using commercially available open-pollinated seed. Pinus radiata is the most widely planted commercial conifer species in the Southern Hemisphere. Reports on transformation of this species have relied on particle bombardment of embryogenic callus derived from immature embryos. The main drawback to the method is the small number of genotypes that are amenable to transformation and regeneration. Since more than 80% of genotypes of radiata pine can be regenerated using cotyledons from mature seed, cotyledon explants were cocultivated with A. tumefaciens strain AGL1 containing a plasmid coding for the neomycin phosphotransferase II (nptII) gene and the -glucuronidase (GUS) gene (uidA). Transformed shoots were selected using either geneticin or kanamycin. Critical factors for successful transformation were survival of the cotyledons after cocultivation and selection parameters. Of the 105 putative transformants that were recovered from selection media, 70% were positive for integration of the nptII gene when analysed by PCR. GUS histochemical assay for uidA expression was unreliable because of reaction inhibition by unidentified compounds in the pine needles. Further, only 4 of the 26 independent transformants characterised by PCR and Southern analysis contained an intact copy of both genes. The remaining 22 transformants appeared to have a truncated or rearranged copy of the T-DNA. It is possible that the truncation/rearrangements are due to the Cauliflower mosaic virus (CaMV) 35S promoter. Analysis of the T-DNA junction sites and sequencing of the introduced DNA will help elucidate the nature of T-DNA insertion so that genetic modification of radiata pine can be targeted effectively.Communicated by P. Debergh 相似文献
9.
In developing mustard (Sinapis alba L.) seedlings continuousred light acting via the agency of phytochrome stimulates therate of expansion of cotyledons. Although phytochrome actionon cotyledon expansion is evident only after 36 h from sowing,the photoresponse escapes from reversibility at about 15 h fromsowing. The time lag of 21 h between loss of photoreversibilityand the onset of photoregulated cotyledon expansion indicatesthe existence of long-lived components in the phytochrome-triggeredsignal chain. Phytochrome-regulated cotyledon expansion doesnot require the involvement of photosynthesis, as applicationof SAN 9789, an inhibitor of chloroplast biogenesis, did notaffect cotyledon expansion. The role of turgor pressure-relatedcellular parameters such as osmotic potential () cell wall extensibility(m), hydraulic conductivity (L) and yield threshold (Y) forcell expansion were examined during photoregulated cotyledonexpansion. Using the general equation of cell growth dv/dt =[Lm/(L+m)]( - Y), where dv/dt is the rate of volumetric growth,it was demonstrated that the light-mediated cotyledon expansionresults from an increase in cell wall extensibility (m). Theseresults are discussed in relation to the photoregulation ofcotyledon expansion. Key words: Cell wall extensibility, growth, Sinapis alba L., phytochrome 相似文献
10.
Agrobacterium -mediated transformation of embryogenic cell suspensions of the banana cultivar Rasthali (AAB) 总被引:9,自引:0,他引:9
T. R. Ganapathi N. S. Higgs P. J. Balint-Kurti C. J. Arntzen G. D. May J. M. Van Eck 《Plant cell reports》2001,20(2):157-162
A protocol was developed for establishing embryogenic suspension cultures from in vitro-grown, thin shoot-tip sections of
the banana cultivar Rasthali. The best medium for callus induction was an MS-based medium supplemented with 2 mg/l 2,4-D and
0.2 mg/l zeatin. The callus was transferred to liquid medium to establish embryogenic cell suspensions. These cultures were
subsequently used for Agrobacterium-mediated transformation. The Agrobacterium tumefaciens strain EHA105 containing the binary vector pVGSUN with the als gene as a selectable marker and an intron-containing the gusA gene as a reporter gene was used for transformations. The herbicide Glean was used as a selection agent. Two hundred putative
transformants were recovered, of which a set of 16 was tested by histochemical analysis for GUS expression and by Southern
blot analysis with a probe for the gusA gene. The plants were positive for GUS expression and integration of the gusA gene. Two of the transformants were grown to maturity under greenhouse conditions. Bananas were harvested to test GUS expression
by histochemical analysis. The fruit from both transgenics tested positive for GUS expression.
Received: 22 February 2000 / Revision received: 2 October 2000 / Accepted: 5 October 2000 相似文献
11.
Factors affecting transient gene expression in cultured radiata pine cotyledons following particle bombardment 总被引:2,自引:0,他引:2
Manuel Rey María Victoria González Ricardo J. Ordás Raffaela Tavazza Giorgio Ancora 《Physiologia plantarum》1996,96(4):630-636
Transfer and expression of the β–glucuronidase gene ( gusA ) in cultured cotyledons of radiata pine ( Pinus radiata D. Don ) were obtained by particle bombardment. Conditions for optimum transient expression were established by using plasmid pB[/12], delivered by gold particles, 1.6 μm in diameter, into 8-day-old cultured cotyledons. Helium pressure of 7.6 MPa, bombardment distance between the stopping screen and the target tissues of 6 cm, and 0.8 μg of plasmid DNA per bombardment proved to be the best parameters for transient expression; using these parameters 79% of bombarded cotyledons showed GUS activity, with 4.3 blue spots per cotyledon. This system was used for studying the expression of several gus-driven promoters the expression of the sunflower ubiquitin gene promoter was higher (99% of positive cotyledons, with 14.2 blue spots per cotyledon) than that of the CaMV 35S promoter, whereas the rice actin and the maize alcohol dehydrogenase gene promoters gave lower gusA expression, as determined histochemically. These results were confirmed by using the gus fluorometric assay. Use of the sunflower ubiquitin gene promoter resulted in gusA expression up to 20 days after bombardment, with a significant level of gus expressing loci per bombarded cotyledon, whereas with the CaMV 35S promoter gusA expression was lost 12 days after bombardment. 相似文献
12.
Y. X. Duan W. W. Guo H. J. Meng N. G. Tao D. D. Li X. X. Deng 《Biologia Plantarum》2007,51(2):212-216
Transformation and high efficient regeneration of transgenic plants from embryogenic calluses of Bingtang sweet orange [Citrus sinensis (L.) Osbeck] was reported. Embryogenic calluses were inoculated with Agrobacterium tumefaciens strain EHA105, harboring the binary Ti plasmid pROK II and carrying a neomycin phosphotransferase II (NPTII) gene, an intron
β-glucuronidase (GUS) gene and the Arabidopsis APETALA1 (AP1) gene. Transformation treatment was with inoculation time of 30 min, co-culture of 3 d at 23 °C and supplementation
of the co-culture medium with 2 mg dm−3 acetosyringone (AS). Kanamycin (50 mg dm−3) was effective to inhibit the growth of non-transformed calluses while it did not affect the transformed ones. The total
number of transformed callus lines was 7 with 100 % embryo induction. High efficient regeneration of the transgenic embryos
(88 % with 4–5 shoots per embryoid) was realized within 3 months. Integration of the transgene into the citrus genome was
confirmed by histochemical GUS staining, polymerase chain reaction (PCR) analysis with AP1-specific primer and Southern blot
hybridization with a 712 bp PCR fragment of AP1 as the probe. 相似文献
13.
Agrobacterium tumefaciens -mediated transformation of soybean [Glycine max (L.) Merrill. cv. Jack] using immature zygotic cotyledons was investigated to identify important factors that affected transformation
efficiency and resulted in the production of transgenic soybean somatic embryos. The factors evaluated were initial immature
zygotic cotyledon size, Agrobacterium concentration during inoculation and co-culture and the selection regime. Our results showed that 8- to 10-mm zygotic cotyledons
exhibited a higher transformation rate, as indicated by transient GUS gene expression, whereas the smaller zygotic cotyledons,
at less than 5 mm, died shortly after co-cultivation. However, the smaller zygotic cotyledon explants were found to have a
higher embryogenic potential. Analysis of Agrobacterium and immature cotyledon explant interactions involved two Agrobacterium concentrations for the inoculation phase and three co-culture regimes. No differences in explant survival or somatic embyogenic
potential were observed between the two Agrobacterium concentrations tested. Analysis of co-culture regimes revealed that the shorter co-culture times resulted in higher explant
survival and higher somatic embryo production on the explants, whereas the co-culture time of 4 days severely reduced survival
of the cotyledon explants and lowered their embryogenic potential. Analysis of selection regimes revealed that direct placement
of cotyledon explants on hygromycin 25 mg/l was detrimental to explant survival, whereas 10 mg/l gave continued growth and
subsequent somatic embryo development and plant regeneration. The overall transformation frequency in these experiments, from
initial explant to whole plant, was 0.03 %. Three fertile soybean plants were obtained during the course of these experiments.
Enzymatic GUS assays and Southern blot hybridizations confirmed the integration of T-DNA and expression of the GUS-intron
gene in the three primary transformants. Analysis of 48 progeny revealed that three copies of the transgene were inherited
as a single Mendelian locus.
Received: 6 December 1999 / Revised: 11 February 2000 / Accepted: 14 March 2000 相似文献
14.
Phytosulphokine-{alpha}, a peptidyl plant growth factor, stimulates somatic embryogenesis in carrot 总被引:1,自引:0,他引:1
Kobayashi T; Eun C; Hanai H; Matsubayashi Y; Sakagami Y; Kamada H 《Journal of experimental botany》1999,50(336):1123-1128
Phytosulphokine- (PSK-) is the first chemically characterized peptide
that acts as a plant growth factor. It stimulates the proliferation of
asparagus and rice cells, but no information is yet available on its
effects on plant morphogenesis. The effects of PSK- on somatic
embryogenesis in carrot (Daucus carota L.) were
examined. PSK-, when added to the induction medium for somatic
embryogenesis, increased the number of somatic embryos. The chemical
analogues [2-5]PSK- and tyrosine sulphate ester (Tyr-SO3 H), which
have been used as negative controls in other systems, had no effect.
Moreover the proliferation of cells during somatic embryogenesis was also
enhanced by PSK- these results indicate that PSK-
enhanced cell division and, as a consequence, stimulated carrot somatic
embryogenesis. PSK- also stimulated the proliferation of
embryogenic cells in medium that contained 2,4-dichlorophenoxyacetic acid
(2,4-D), in which somatic embryos did not form, as well as the
proliferation of non-embryogenic cells (cells that had lost the ability to
form somatic embryos) in medium without 2,4-D. These results indicate that
PSK- has a stimulatory effect on cell division generally in carrot
cell cultures.Key words: Daucus carota, plant growth
factor, somatic embryogenesis, sulphated peptide.
相似文献
15.
High-light damage in air-dry thalli of Lobaria
pulmonaria were measured in the laboratory as reductions in
maximal PSII efficiency (Fv/Fm)
after a 48 h recovery in a hydrated state at low light to account for
permanent damage. Thalli treated with the lowest light dose (90 mol photons
m-2) recovered normal
Fv/Fm-values with increasing irradiances (400-700 nm)
up to 1000 mol photons
m-2 s-1. Doubling this dose
lowered the threshold level for damage from 1000 to 320 mol photons m-2
s-1, and reduced
Fv/Fm at 1000 mol
photons m-2 s-1 by more than
50%. A second doubling of the dose to 360 mol photons
m-2 caused damage at 200 mol photons
m-2 s-1, and a nearly
complete cessation of PSII efficiency occurred at 1000 mol photons
m-2 s-1. No reciprocity of
irradiance and duration of illumination for PSII function was found. The
measured time-dependent decrease in
Fv/Fm was remarkably similar for
the naturally coupled, but artificially separated, light and temperature
factors. Therefore, the damage of high light on desiccated L.
pulmonaria seemed to be an additive effect of high irradiance
and high temperatures. Air-dry thalli were highly heat susceptible, being
affected already at temperatures around 40C.
Logging operations in forests are likely to raise the solar radiation at
remaining lichen sites to destructive levels.Keywords:
Lichens, high-light damage, heat stress, poikilohydric
organisms, reciprocity.
相似文献
16.
The involvement of Na+, K+,
Cl- or Ca2+ in the regulation
of salinity stress-induced proline accumulation via the inhibition of the
activity of proline dehydrogenase (PDH; EC 1.4.3.1), a catabolic enzyme of
proline, was investigated in the marine green macroalga Ulva
fasciata Delile. After 6 h of exposure to elevated artificial
seawater (ASW) salinity, adjusted either by increasing the NaCl content in
30 ASW (a change in ion ratio) or by concentrating ASW (a
constant ion ratio), the contents of Na+,
K+ and Cl- linearly
accumulated with increasing salinity from 30-90 (parts per
thousand); the accumulation pattern of each ion was similar between the two
treatments. An increase in NaCl content in ASW induced proline
accumulation, but decreased both the PDH activity and the total
water-soluble Ca2+ contents, while concentrated ASW
had no effect. As compared to a constant value at 30, both the
contents of total and water-soluble CA2+ and the
activity of PDH decreased 1 h after exposure to 60 (adjusted by
increasing NaCl content in 30 ASW) and concomitantly the
content of seawater Ca2+ increased, while proline
accumulated after 3 h. The addition of 15 mM ethylene
glycol-bis-(2-aminoethyl ether)
N,N,N-tetraacetic acid (EGTA) in
60 ASW (adjusted by increasing the NaCl content in
30 ASW) enhanced both the proline accumulation and the decrease
in the content of total and water-soluble cellular
Ca2+ and the activity of PDH; the effects of EGTA
were reversed by 10 mM CaSO4. These results indicate that a loss of
cellular Ca2+ is associated with the NaCl induction
of proline accumulation via an inhibition of PDH activity in U.
fasciata. 相似文献
17.
A karyopherin (LeKAP1) cDNA was isolated from tomato
plants. The deduced LeKAP1 protein sequence of 527 amino acids
showed similarity to other plant karyopherin proteins. When
LeKAP1 was expressed in a yeast two-hybrid
system together with the gene coding for the capsid protein (CP) of the
tomato yellow curl leaf virus (TYLCV), it interacted directly with CP.
Thus, LeKAP1 may be involved in the nuclear import of TYLCV CP
and, potentially, the TYLCV genomes during viral infection of the host
tomato cells. 相似文献
18.
Mature seed-derived embryogenic calli of indica rice (Oryza sativa L. cv. PAU201) were induced on semisolid Murashige and Skoog medium supplemented with 2.5 mg dm−3 2,4-dichlorophenoxyacetic acid + 0.5 mg dm−3 kinetin + 560 mg dm−3 proline + 30 g dm−3 sucrose + 8 g dm−3 agar. Using OsglyII gene, out of 3180 calli bombarded, 32 plants were regenerated on medium containing hygromycin (30 mg dm−3). Histochemical GUS assay of the hygromycin selected calli revealed GUS expression in 50 % calli. Among the regenerants,
46.87 % were GUS positive. PCR analysis confirmed the presence of the transgene of 1 kb in 60 % of independent plants. Further,
these plants have been grown to maturity in glasshouse. In vitro screening for salt tolerance showed increase in fresh mass of OsglyII putative transgenic calli (185.4 mg) as compared to control calli (84.2 mg) on 90 mM NaCl after 15 d. When exposed to 150
mM NaCl, OsglyII putative transgenic plantlets showed normal growth while the non-transgenic control plantlets turned yellow and finally did
not survive. 相似文献
19.
Eliane R. Santarém John J. Finer 《In vitro cellular & developmental biology. Plant》1999,35(6):451-455
Summary Transgenic soybean can be efficiently produced by particle bombardment of embryogenic suspension culture material. Unfortunately,
the time required to obtain a transformation-competent soybean suspension culture line is often lengthy and can result in
reduced fertility of regenerated plants. In addition, establishment and maintenance of embryogenic suspension cultures can
be very difficult. The objective of this work was to minimize the time required to obtain transformation-competent embryogenic
tissue and optimize DNA delivery into that tissue. Somatic embryos were induced from immature cotyledons of soybean [Glycine max (L.) Merrill cv ‘Jack’] by placement of cotyledons, adaxial side up, on a MS-based induction medium containing 40 mg (181
μM) 2,4-dichlorophenoxyacetic acid (2,4-D) per 1 and 6% sucrose. Embryogenic tissues, which formed from the surface of the cotyledons
within 2–4 wk, were transferred to an embryo proliferation medium containing 20 mg (90 μM) 2,4-D per 1 and 3% sucrose. After 4 wk, proliferative embryogenic tissue could be used for transformation via particle bombardment.
Desiccation of target tissue, period of subculture prior to bombardment, and the number of bombardments per target tissue
were evaluated for enhancement of transient β-glucuronidase (GUS) expression. The highest number of blue foci was observed
when the target tissue was desiccated for 10 min in an uncovered Petri plate containing proliferation medium, subcultured
on the same day of bombardment, and bombarded three times on a single day. For stable transformation, selection was started
20 d after bombardment using 9 mg hygromycin per 1 for 4 wk, and 18 mg per 1 thereafter. Stably transformed clones were obtained
from tissue bombarded once and twice on a single day. GUS assays and Southern hybridization analysis of DNA from putative
clones confirmed stable integration of the introduced genes. Fertile transgenic plants were obtained in 11–12 mo following
culture initiation. 相似文献
20.
Influence of UV-B radiation and Cd2+ on chlorophyll fluorescence, growth and nutrient content in Brassica napus 总被引:8,自引:0,他引:8
The possible interaction of two stresses, UV-B radiation and cadmium,
applied simultaneously, was investigated in Brassica
napus L. cv. Paroll with respect of chlorophyll fluorescence,
growth and uptake of selected elements. Plants were grown in nutrient
solution containing CdCl2, (0, 0.5, 2 or 5 M)
and irradiated with photosynthetically active radiation
(PAR, 400-700 nm, 800 mol m-2
s-1) with or without supplemental ultraviolet-B
radiation (UV-B, 280-320 nm, 15 kJ m-2
d-1, weighted irradiance). After 14 d of treatment,
the most pronounced effects were found at 2 and 5 M CdCl2 with and
without supplemental UV-B radiation. Exposure to cadmium significantly
increased the amount of Cd in both roots and shoots. In addition, increases
occurred in the concentrations of Fe, Zn, Cu, and P in roots, while K was
reduced. In shoots the S content rose significantly both in the presence
and absence of UV-B radiation, while significant increases in Mg, Ca, P,
Cu, and K occurred only in plants exposed to Cd and UV-B radiation.
Manganese decreased significantly under the combined exposure treatment.
The rise in S content may have been due to stimulated glutathione and
phytochelatin synthesis. Cadmium exposure significantly decreased root dry
weight, leaf area, total chlorophyll content, carotenoid content, and the
photochemical quantum yield of photosynthesis. As an estimation of energy
dissipation processes in photosynthesis, non-photochemical quenching
(qNPQ) was measured using a pulse amplitude modulated
fluorometer. The qNPQ increased with increasing Cd,
while the combination of cadmium and UV-B reduced the
qNPQ compared to that in plants exposed only to
cadmium or UV-B radiation. The chlorophyll a:b ratio
showed a reduction with UV-B at no or low Cd concentrations (0 M,
0.5 M CdCl2), but not at the higher Cd concentrations used (2
M, 5 M CdCl2). Thus in some instances there appeared to
be a UV-B and Cd interaction, while in other plants response could be
attributed to either treatment alone.Keywords:
Brassica napus, cadmium, ultraviolet-B
radiation.
相似文献