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  • Nicotiana
  • Springer  (63)
  • PANGAEA
  • Periodicals Archive Online (PAO)
  • 2005-2009
  • 1980-1984  (63)
  • 1925-1929
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  • 1
    ISSN: 1432-2048
    Keywords: mRNA ; Protein synthesis ; Protoplasts ; Nicotiana ; Translation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Studies of proteins synthesized in vitro by messenger RNA (mRNA) extracted from tobacco protoplasts showed that the changes in protein synthesis and especially the lack of certain proteins observed previously in isolated protoplasts did not result from a failure of translation.
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  • 2
    Electronic Resource
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    Planta 148 (1980), S. 462-467 
    ISSN: 1432-2048
    Keywords: Flower formation ; Leaves (in flower formation) ; Meristem (shoot) ; Nicotiana ; Roots (and flower formation)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The terminal, apical shoot meristem ofN. tabacum cv. Wisconsin 38 normally differentiates into a flower after producing 30 to 40 nodes. The influence of leaves and roots on the regulation of flowering was evaluated by counting the number of nodes produced after removal of leaves or the induction of adventitious roots. Leaf removal has no effect on the number of nodes produced before flower formation. Root induction significantly increases the number of nodes produced before flower formation. The plant behaves as if it were measuring the number of nodes between the meristem and the roots as a means of regulating meristem conversion from vegetative to floral differentiation.
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  • 3
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    Planta 148 (1980), S. 491-497 
    ISSN: 1432-2048
    Keywords: Leaves (polysomes) ; Nicotiana ; Polysomes ; Poly(A)+ RNA ; Protein synthesis ; RNA (polysomal, polyA+)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The isolation of intact polysomes from leaves of tobacco (Nicotiana tabacum L.) is dependent on the age and state of development of leaves. Undegraded polysomes from young leaves in the early stages of expansion can be isolated easily by extracting the leaves in ice-cold extraction buffer (200 mM tris(hydroxymethyl)aminomethylmethane(Tris)-HCl, pH 9.0; 400 mM KCl; 200 mM sucrose; 35 mM MgCl2). Medium-size leaves give best yields of undegraded polysomes when extractions are carried out in the above buffer and in the presence of ethyleneglycol-bis-(β-amino-ethyl ether)-N,N′-tetracetic acid (EGTA) and mercaptoethanol. Isolation of polysomes from large, nearly fully expanded (mature) leaves requires all of the above plus diethyldithiocarbamate (DIECA) in the extraction medium. An extraction medium consisting of 25 mM EGTA, 0.01 M mercaptoethanol, 25 mM DIECA and 0.5% of the nonionic detergent, Nonidet-P40 (NP 40) was found to be very suitable for extraction of polysomes from all developmental stages of leaves. The polysomes extracted in the above medium showed active translation of protein in the wheat-germ in-vitro protein-synthesizing system. The translational products were similar when translations were carried out directly with polysomes or polysomal RNA, or polysomal poly(A)+ RNA from tobacco leaves. Poly(A)− polysomal RNA was a poor template in the in-vitro wheat-germ system.
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  • 4
    ISSN: 1432-2048
    Keywords: Auxin metabolism ; Cell suspensions (low density) ; Medium conditioning ; Nicotiana ; Nitrogen metabolism ; Protoplasts ; Vitamin requirements
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Preliminary attempts to define a completely synthetic medium able to support divisions of haploid tobacco mesophyll protoplasts at low initial densities have failed. High protoplast concentrations together with large amounts of naphtaleneacetic acid in the medium (3 mg l-1 NAA) were required for maximal induction of protoplast division. However, cell suspensions derived from haploid protoplasts after four days of preculture at high initial cell densities could be diluted to densities as low as 1–4 cells ml-1, provided the concentration of NAA in the medium was lowered to below 0.3 mg l-1. The optimal NAA supply for low cell density growth was affected by the nature of the nitrogen source. A simple minimal medium which supports the growth of these haploid cells with a plating efficiency of 30–40%, independent of the cell density in the range of 1–4 to 3·104 cells ml-1, has been established. In this medium inositol was the only vitamin stringently required for growth. Growth of cells at low densities was also possible in a medium initially containing 3 mg l-1 NAA, provided it was conditioned by the growth of protoplasts at high densities. Preliminary experiments with [14C]NAA showed that the amount of free NAA remaining in the medium after preincubation at high densities was drastically reduced. Simultaneously, NAA conjugates accumulated in the medium. The implications of these results are discussed.
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  • 5
    ISSN: 1432-2048
    Keywords: Auxin receptor ; Callus ; Nicotiana
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Cultured tobacco-pith tissue contains a cytoplasmic receptor for indoleacetic acid (IAA). The concentration of binding sites is very low in comparison to that of several auxin receptors found by other investigators. A few obvious possible causes (degradation or inactivation) were investigated. From the results we conclude that the low number of binding sites is real. The receptor binds IAA optimally at pH 7.5–7.8 and at a temperature of 24–30°C, when incubated for 25–30 min. The binding is very specific, as is shown by competition experiments. The concentration of the receptor in the callus tissue changes dramatically during each culture period, which suggests a possible role in development. The receptor was partly purified by gel filtration on Sepharose 6B followed by ion-exchange chromatography on DEAE-cellulose.
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  • 6
    ISSN: 1432-2048
    Keywords: 1-Aminocyclopropane-1-carboxylic acid ; Avena ; Ethylene ; Etiolation ; Light (ethylene production) ; Nicotiana
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract White light inhibits the conversion of 1-amino-cyclopropane-1-carboxylic acid (ACC) in discs of green leaves of tobacco (Nicotiana tabacum L.) and segments of oat (Avena sativa L.) leaves by from 60 to 90%. Etiolated oat leaves do not show this effect. The general nature of the effect is shown by its presence in both a mono- and a dicotyledon. Since the leaves have been grown and pre-incubated in light, yet can produce from 2 to 9 times as much ethylene in the dark as in the light, it follows that the light inhibition is fully reversible. The inhibition by light is about equal to that exerted in the dark by CoCl2; it can be partly reversed by dithiothreitol and completely by mercaptoethanol. Thus the light is probably acting, via the photosynthetic system, on the SH group(s) of the enzyme system converting ACC to ethylene.
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  • 7
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    Planta 149 (1980), S. 205-206 
    ISSN: 1432-2048
    Keywords: Callus cultures ; Nicotiana ; Nicotine ; Tissue cultures
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Callus cultures of two low-alkaloid lines of Nicotiana tabacum L. had considerably lower nicotine contents than cultures from the respective highalkaloid cultivars which were isogenic except for the two loci for alkaloid accumulation. Thus, there was a strong correlation between the nicotine content of callus cultures and the plants from which they were derived.
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  • 8
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    Planta 150 (1980), S. 9-12 
    ISSN: 1432-2048
    Keywords: Auxin binding ; Nicotiana ; Plasma membrane ; Protoplast
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In vitro binding of 1-naphthaleneacetic acid (NAA) to particulate fractions from tobacco leaf protoplasts was studied. In freshly isolated protoplasts no specific binding could be detected, whereas it was present in particulate fractions from tobacco leaves. It is concluded that the NAA-binding-sites are probably located at the external face of the plasma membrane; they are destroyed during protoplast isolation by proteolytic enzymes in the cellulase and macerozyme preparations. After culturing the protoplasts for 3–4 d, the first cell divisions were observed and at the same time specific NAA-binding became detectable. The affinity constant for NAA was approx. 2·106 mol-1 and the number of binding sites increased during further culture.
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  • 9
    ISSN: 1432-2048
    Keywords: Cytokinin habituation ; Habituation, cytokinin ; Nicotiana ; Position effects (habituation) ; Size effects (habituation) ; Tissue culture
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Pith tissue of Nicotiana tabacum L. cv. “Havana 425” exhibits a gradient in its tendency to habituate for cytokinin on an auxin-containing medium at 35° C, about 10° C above the standard culture temperature. Explants of pith from below the 8th to 11th internode, counting from the bottom of the plant, rarely habituate for cytokinin; explants from above this threshold habituate rapidly. The explants must also be above a critical size, about 20–30 mg, to habituate. There was a pronounced interaction between size and position effects; the threshold position for cytokinin habituation shifted upward with decreasing explant size.
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  • 10
    ISSN: 1432-2048
    Keywords: Abscisic acid ; Nicotiana ; Ribonucleic acid synthesis ; Tobacco mosaic virus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Uptake of abscisic acid from the culture medium by discs of healthy and tobacco mosaic virus-infected tobacco leaves was measured. Small (two to five-fold) increases in abscisic acid concentration in discs caused increases in rates of [3H]uridine and [3H]adenine incorporation into total nucleic acid, virus RNA and host ribosomal RNA. Net accumulation of virus RNA was also enhanced by abscisic acid. This evidence for stimulation of RNA synthesis is compared with previous reports showing inhibition of RNA synthesis in other tissues. It is suggested that the increase in endogenous abscisic acid caused by tobacco mosaic virus infection may be at least partly responsible for observed increases in rates of RNA synthesis after infection.
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  • 11
    ISSN: 1432-2048
    Keywords: Aurea mutant ; Chlorophyll-protein complexes ; Chloroplast grana ; Light-harvesting pigment-protein complex ; Nicotiana
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Phenotypic characterizations of the semidominant aurea tobacco (Nicotiana tabacum L.) mutant Su/su, the homozygous mutant Su/Su and three green revertants (R1, R2, and R3) are presented. The leaf color of Su/su plants varies from yellow to light-green when grown under high and low energy fluence rates (33.0 and 3.3 W m−2), respectively. The change in visual phenotype under high-light conditions is correlated with decreased content of chlorophyll per leaf area, agranal chloroplast ultrastructure, changes in the number of chlorophyll-protein complexes, and absence of two or more of the light harvesting chlorophyll-polypeptides of 25,000–29,000 dalton. The homozygous mutant grown under low light was shown to be completely lacking in grana stacks and to be deficient in chlorophyll-protein complexes. Revertant R1 was found to be identical to wild-type plants in all parameters examined (leaf color, chloroplast ultrastructure, chlorophyll-protein complexes, chlorophyll-protein complex polypeptides) except in chlorophyll content. It did not show an increased chlorophyll and carotenoid content as did the wild-type plants when exposed to high light. Revertants R2 and R3 were similar to the heterozygous mutant Su/su in most of the parameters examined. They yellowed because of a loss of chlorophyll and an increase in the amount of carotenoids, had agranal chloroplasts, and had variant chlorophyll-protein complexes when grown under high light intensities. However, each appeared to contain some of the light-harvesting pigment-protein complex polypeptides found to be absent in Su/su when grown under high-light conditions.
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  • 12
    ISSN: 1432-2048
    Keywords: Chloroplast ; Cybrid ; Nicotiana ; Protoplast ; Somatic hybrid ; Sterility, male
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Protoplasts from a nitrate reductase-deficient mutant of Nicotiana tabacum L. were fused with protoplasts from a stamen-less, cytoplasmically malesterile cultivar of tobacco containing the cytoplasm from N. suaveolens Lehm. Plants were regenerated from the fused protoplasts and characterized with respect to stamen development, chromosome number, and chloroplast composition. Of 29 regenerated plants, stamen production was restored in 26 plants and pollen production in 22. One plant was male sterile and two plants have never flowered. Analysis of the electrophoretic mobility of ribulose-1,5-bisphosphate carboxylase (RuBPcase) showed that 19 of the plants contained RuBPcase of the N. suaveolens type, six plants contained enzyme of the N. tabacum type, and four plants contained both types. Analysis of resistance to tentoxin in seedlings from 20 of the plants demonstrated that 14 had N. suaveolens-type chloroplasts, three had N. tabacum type, and three contained both types. Many of the plants which produced stamens and pollen still contained chloroplasts of the N. suaveolens type. Thus, the trait of cytoplasmic male sterility in tobacco is not an expression of the type of chloroplast genetic material.
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  • 13
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    Planta 154 (1982), S. 241-250 
    ISSN: 1432-2048
    Keywords: Callose ; Cellulose ; Nicotiana ; Pectin ; Pollen tube wall
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Tobacco pollen tubes grown in vitro and from pollinated tobacco styles were treated by chemical solvents to remove one or more of the following polysaccharides from the tube walls: pectin (ethylenediamine tetraacetic acid); hemicellulose (alkali); callose (alkali; potassium hypochlorite); cellulose (cuprammonium); and all polysaccharides with exception of cellulose (H2O2/glacial acetic acid). Both the inner tube wall, which we had regarded as the secondary wall, and the plugs contained, in addition to callose, microfibrils of cellulose and “non-cellulosic” microfibrils that had “pectin-like” properties. When using the expressions callosic or callose layer and callose plugs in reference to pollen tubes, one should realize that they do not imply the exclusive presence of callose in the inner tube wall layer and its localized thickenings.
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  • 14
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    Planta 153 (1981), S. 42-48 
    ISSN: 1432-2048
    Keywords: Crown gall ; Hyperplasia ; Hypertrophy ; Teratoma ; Nicotiana
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Crown-gall teratomas are tumors of higher plants with an intrinsic capacity for organogenesis. The growth pattern of tobacco (Nicotiana tabacum L.) teratoma shoots, which is highly aberrant in primary tumors, becomes normal when the shoots are grafted to healthy stock plants. However, certain abnormalities commonly persist; tumors form at the graft junctions, leaves are small, apical dominance is incomplete, the stem and proximal region of the leaf midribs swell excessively, and localized eruptions of neoplastic growth occur on the swollen tissue. Swelling of the shoots is primarily the result of cell hypertrophy in the cortex. Neoplastic divisions do not occur as a general rule; they are restricted, with the exception of tumor formation at the graft junctions, to localized eruptions of teratoid growth on the nodes and leaf midribs where cell hypertrophy is most evident. The histology of the apical meristem and histogenesis of primary tissues is normal, even in grossly distorted shoots. Similarly, there is no evidence of unregulated division in the vascular cambium. It is concluded that cell expansion and division are tightly regulated in meristematic regions of teratoma shoots whereas post-meristematic tissue is prone to excessive hypertrophy and eventual initiation of neoplastic cell division.
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  • 15
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    Planta 153 (1981), S. 511-518 
    ISSN: 1432-2048
    Keywords: Amino-acid transport ; Arginine ; Cell culture (tobacco) ; Nicotiana ; Transport (arginine)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Arginine transport in suspension-cultured cells of Nicotiana tabacum L. cv. Wisconsin-38 was investigated. Cells that were preincubated in the presence of Ca2+ for 6 h prior to transport exhibited stimulated transport rates. After the preincubation treatment, initial rates of uptake were constant for at least 45 min. Arginine accumulated in the cells against a concentration gradient; this accumulation was not the result of exchange diffusion. Arginine uptake over a concentration range of 2.5 μM to 1 mM was characterized by simple Michaelis-Menten kinetics with a Km of 0.1 mM and a Vmax of 9,000 nmol g-1 fresh weight h-1. Transport was inhibited by several compounds including carbonylcyanide-m-chlorophenylhydrazone, 2,4-dinitrophenol, N,N′-dicyclohexylcarbodiimide, and N-ethylmaleimide. Inhibition by these compounds was not the result of increased efflux resulting from membrane damage. A variety of amino acids and analogs, with the exception of D-arginine, inhibited transport, indicating that arginine transport was mediated by a general L-aminoacid permease. Competition experiments indicated that arginine and lysine exhibited cross-competition for transport, with Ki values similar to respective Km values. Arginine transport and low-affinity lysine transport are probably mediated by the same system in these cells.
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  • 16
    ISSN: 1432-2048
    Keywords: Cell suspension culture ; p-Fluorophenylalanine (metabolism of) ; Nicotiana ; Phenylalanine ammonia-lyase ; Resistant cell
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The metabolism of D- and L-p-fluorophenylalanine (PFP) in DL-PFP resistant and sensitive tobacco cell cultures (Nicotiana tabacum), cell lines TX4 and TX1, respectively, has been compared. The amino acid analogue was taken up at a lower rate by the resistant cell line TX4. Incorporation of PFP into protein was also considerably reduced in TX4 cells, compared to TX1 cells. This, however, resulted mainly from a diminished availability of PFP due to a more rapid conversion of PFP by TX4 cells. TX1 cells and TX4 cells converted PFP qualitatively in the same way. The only detectable metabolite of D-PFP was N-malonyl-D-PFP, while all metabolites of L-PFP were identified as sequent products of the initial deamination of L-PFP by the enzyme phenylalanine ammonia-lyase (PAL). As TX4 cells were endowed with higher PAL-activity than TX1 cells, the resistant cells were able to metabolize L-PFP more rapidly to give, e.g., p-fluorocinnamoyl glucose ester and p-fluorocinnamoyl putrescine. In the presence of the specific PAL-inhibitor α-aminooxy-β-phenylpropionic acid TX4 cells were slightly more sensitive to PFP. This suggests that the better “detoxification” contributes to the acquired resistance. The use of PFP as specific indicator for cell lines with increased PAL-activity, and hence increased levels of phenolic compounds, is discussed.
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  • 17
    ISSN: 1432-2048
    Keywords: Cell differentiation ; Chromosomal protein ; Nicotiana ; Protein, non-histone ; Template activity ; Tissue culture (chromatine)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Non-histone chromosomal proteins (NHP) were isolated from different stages of Nicotiana tabacum L. pith dedifferentiation to callus and callus redifferentiation. The NHP were separated by sodium dodecylsulfate-polyacrylamide gel electrophoresis on slab gels and analyzed by densitometry. Simultaneous histological changes are reported. In both processes, some high molecular weight protein (HMWP) bands increase drastically in an induction period, previous to cell proliferation, and decrease when cell division declines. Some low molecular weight protein bands, intense in pith tissue, decrease early when callus is forming and increase when cells differentiate. chromatin template activity is high when cells proliferate, coinciding with maximum HMWP-bands intensity.
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  • 18
    ISSN: 1432-2048
    Keywords: Floral induction ; Nicotiana ; Pollen embryogenesis ; Pollen sterility ; Sex expression
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Pollen sterility, sex balance, and floral induction of the pollen donor plants were tested for a possible relation to embryogenesis from in vitro cultured tobacco pollen (Nicotiana tabacum L. var. Badischer Burley). The pollen grains destined to become embryos in culture (P-grains) were sterile for the donor plants as judged by their staining reaction with acetocarmine and fluorescin-diacetate, and by an in vitro germination test. They were produced in high frequency in flowers which exhibited a shift in sex balance towards femaleness. Sex balance could be measured by the relative length of pistil to stamens. High P-grain frequency, high pollen sterility, and a shift in sex balance towards femaleness could be induced by raising the donor plants under short days and/or low temperature (18–15° C) as compared to long days at 24° C. Short days and/or low temperature also reinforced floral induction, revealing that the tobacco variety Badischer Burley is a quantitative short day and low temperature plant and that the variety follows the rule that conditions of strong floral induction shift sex balance towards femaleness. At 12° C and short days, contabescent flowers were formed with completely sterile anthers containing a few and mostly collapsed P-grains. Based on these results, it is now possible to predict conditions by which haploids via pollen embryogenesis might be produced in high frequency from low-yielding and recalcitrant species.
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  • 19
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    Planta 156 (1982), S. 402-406 
    ISSN: 1432-2048
    Keywords: Alternation of generations ; Apogamy ; Nicotiana ; Pollen embryogenesis ; Sexuality
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The previously-found correlation of P-grain frequency in situ (pollen grains which are able to form embryos in culture) with floral induction and sex balance of tobacco plants (Nicotiana tabacum L. var. Badischer Burley) was studied in more detail in order to find out in which stage of sexual development P-grain induction takes place. To know the time of P-grain induction is important for attempts to intervene more specifically in plant development, particularly with chemicals, with the aim of inducing P-grain formation. It was found that the floral stimulus was not involved in the control of both sex balance and P-grain formation. Rather, sex balance and P-grain formation were controlled by factors operating during flower development, that is, after floral induction period. Furthermore, both phenomena seemed to be controlled by the same factors, since changes in P-grain frequency and sex balance followed the same time course when flowering plants were transferred from short-day conditions at 24° C to 18° C and vice versa. These transfer experiments also revealed that the process of P-grain induction starts early in flower development, that is, well before meiosis (about five weeks before anthesis), but that the potential P-grains can return to normal gametophytic development until after meiosis. Pollen embryogenesis is regarded as a form of induced apogamy and is discussed in relation to an alternation of generations.
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  • 20
    ISSN: 1432-2048
    Keywords: Hydrogenperoxide ; Lignification ; Nicotiana ; Peroxidase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Three peroxidase isoenzyme-groups found in cell walls of tobacco were tested for their capacity to form H2O2. Isoenzyme-group GI, located only in cell walls (GII and GIII are also found in protoplasts) showed the highest Kapp-value for H2O2-formation. The lowest Kapp-value, i.e., maximal H2O2-formation was received for group GIII which is ionically bound to the cell wall. As shown before, GI yields maximal polymerization rates for coniferyl- and p-coumarylalcohol. These facts indicate that each of the peroxidase isoenzyme groups of the cell wall is involved with different catalytic functions within the same pathways of H2O2-formation and succeeding lignification. H2O2-formation catalyzed by all 3 groups was increased by very low concentrations of Mn2+-ions. The required amount of Mn2+ leading to maximal stimulation was in each case dependent on the basic rate of H2O2-formation. Maximal stimulation of H2O2-formation by phenolic compounds was achieved by coniferylalcohol at a concentration of 10-4M for all groups. Stimulation by p-coumaryl-and by sinapylalcohol was not as significant.
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  • 21
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    Planta 149 (1980), S. 402-407 
    ISSN: 1432-2048
    Keywords: Cytokinin ; Habituation rates ; Nicotiana ; Temperature and habituation ; Tissue culture
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Pith parenchyma tissue ofNicotiana tabacum L. cv. “Havana 425” becomes cytokinin habituated when incubated at 35°C on an auxin-containing medium. Under these conditions, habituated, hyperplastic nodules appear on the tissues. We used these nodules to estimate the incidence of habituation by a statistical method. The rate of habituation varied with the season. Tissue isolated from plants in the spring habituated approx. 7 times faster than did tissue isolated from plants in winter. The fact that the average rate, 〉4×10−3 per cell generation, was 100–1,000 times faster than the rate of somatic mutation inNicotiana species and depended on the physiological state of the tissue provides further evidence that habituation involves epigenetic changes rather than rare, random genetic mutations. We also found that kinetin (6-furfurylaminopurine) induced habituation and that the concentration required depended on the duration of cytokinin treatment. For long incubation times, approx. 6×10−10 M kinetin, which is about 1,000-fold lower than the concentration optimal for growth of cytokinin-requiring pith tissue, was sufficient to induce habituation. These results support the hypothesis that the habituated state is maintained by a positive feedback loop in which cytokinins either induce their own synthesis or inhibit their own degradation.
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  • 22
    ISSN: 1432-2048
    Keywords: Acetate incorporation ; Catharanthus ; Cell suspension cultures ; Fatty acids ; Glycihe ; Nicotiana ; Temperature and fatty-acid synthesis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The fatty-acid composition of C. roseus and N. tabacum cell suspension cultures was unaffected by subculture on Wood and Braun, Murashige and Skoog, or Gamborg B5C media. However, placing the cultures — which were normally grown at 25° C — at 15° C reduced growth but resulted in enhanced formation of oleic and linolenic acids in C. roseus cultures and increased levels of linoleic and linolenic acids in cultures of G. max and N. tabacum, respectively. The incorporation of [14C]acetate into [14C]linoleic acid was more rapid in N. tabacum cells than in G. max cells, but was very poor in C. roseus where the [14C] label was distributed mainly between palmitic and oleic acids.
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  • 23
    ISSN: 1432-2048
    Keywords: Fraction-1 protein ; Hybrid, somatic ; Nicotiana ; Protein (Fraction 1) ; Ribulose-1,5-bisphosphate carboxylase ; Somatic hybrid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract In the sexual interspecific cross, Nicotiana rustica L.xN. tabacum L., N. rustica can serve as the female but not as the male parent. By fusion of protoplasts, the barrier to fertilization was overcome and somatic hybrids containing N. tabacum cytoplasm were produced as shown by isoelectric focusing of the Fraction-1 protein (F-1-protein). All somatic hybrids displayed polypeptides of the large subunit of F-1 protein (which is coded by the chloroplast genome) characteristic of only one or the other parental species. Two hybrids had large subunits of the N. tabacum type and two hybrids had those of the N. rustica type. Three hybrids contained three smallsubunit polypeptides (coded by the nuclear genome), one being characteristic of N. rustica, one characteristic of N. tabacum, and one with an isoelectric point common to both species. A fourth hybrid contained only two small-subunit polypeptides of the N. tabacum type but in a F-1 protein macromolecule whose large subunits were of the N. rustica type. One somatic hybrid was self-fertile and its F2 progeny contained large- and small-subunit polypeptides indistinguishable in their isoelectric points from those in the parent F1 hybrid. All somatic hybrids showed an aneuploid chromosome number and morphological characteristics intermediate between those of N. rustica and N. tabacum.
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  • 24
    ISSN: 1432-2048
    Keywords: Catharanthus ; Cell suspension cultures ; Glycine ; Fatty acids (synthesis, desaturation) ; Nicotiana ; Linoleic, linolenic, oleic, acids ; Temperature and fatty acids
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Cell suspension cultures of Catharanthus roseus G. Don, Glycine max (L.) Merr. and Nicotiana tabacum L. were incubated with [14C]acetate, [14C]oleic acid and [14C]linoleic acid at five different temperatures ranging from 15 to 35° C. When the incubation temperature was increased, [14C]acetate was incorporated preferentially into [14C]palmitate, with a concomitant drop in [14C]oleate formation. Between 15 and 20° C, [14C]oleic acid accumulated in C. roseus cells. In all cultures, optimum desaturation of [14C]oleic acid to [14C]linoleic acid occurred between 20 and 25° C, and in G. max this was also the optimal range for desaturation of [14C]linoleic acid to [14C]linolenic acid. Elongation of [14C]palmitic acid was inhibited when cultures grown at 15° C for 25 h were subsequently incubated with [14C]acetate at 25° C. [14C]oleic acid accumulated in G. max and C. roseus cultures grown at 35° C for 25 h and subsequently incubated at 25° C. Desaturation of [14C]oleic acid increased up to 25° C, but then decreased or leveled off depending on the cell line and on the temperature prior to incubation.
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  • 25
    ISSN: 1432-2048
    Keywords: Chloroplast transfer ; Nicotiana ; Protoplasts fusion and inactivation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Protoplasts of a light sensitive plastome mutant of Nicotiana tabacum (2 n=48) were irradiated and fused with iodoacetate-treated Nicotiana plumbaginifolia (2 n=20) protoplasts. Treated parental protoplasts were unable to divide. Metabolic complementation, however, helped the recovery of interspecific fusion products which survived and formed calli. Altogether 40 clones were investigated. N. plumbaginifolia plants were obtained in 15 clones (38%), somatic hybrids in 23 clones, and both types of regenerates were found in 2 clones. Irradiation therefore significantly increased the frequency of segregant formation with the non-irradiated N. plumbaginifolia nuclei (the frequency was 1.4% in the absence of irradiation). Regenerated plants in most cases (31 out of 34) contained chloroplasts from the irradiated parent. In 6 clones plants were obtained with both types of chloroplast. Thus, irradiated N. tabacum chloroplasts had an improved chance of dominating the heterokaryonderived cells, many of which contained N. plumbaginifolia nucleus. The system described should be generally applicable for the transfer of chloroplasts without the use of selectable genetic markers.
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  • 26
    ISSN: 1432-2048
    Keywords: Auxin-binding site ; Cell suspension culture ; Growth cycle ; Nicotiana
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract We studied the modulation of the number of membrane-bound naphthaleneacetic acid (NAA)-binding sites during the growth cycle of tobacco cells in batch culture. Both cell number and specific NAA-binding increased exponentially, but at different rates and for different periods. This caused a characteristic modulation of the number of binding sites per cell during the growth cycle: During the first day of the lag phase this number decreased; in the exponential phase it rose markedly, and in the stationary phase it was constant.
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  • 27
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    Planta 148 (1980), S. 491-497 
    ISSN: 1432-2048
    Keywords: Leaves (polysomes) ; Nicotiana ; Polysomes ; Poly(A)+ RNA ; Protein synthesis ; RNA (polysomal, polyA+)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The isolation of intact polysomes from leaves of tobacco (Nicotiana tabacum L.) is dependent on the age and state of development of leaves. Undegraded polysomes from young leaves in the early stages of expansion can be isolated easily by extracting the leaves in ice-cold extraction buffer (200 mM tris(hydroxymethyl)aminomethylmethane(Tris)-HCl, pH 9.0; 400 mM KCl; 200 mM sucrose; 35 mM MgCl2). Medium-size leaves give best yields of undegraded polysomes when extractions are carried out in the above buffer and in the presence of ethyleneglycol-bis-(β-amino-ethyl ether)-N,N′-tetracetic acid (EGTA) and mercaptoethanol. Isolation of polysomes from large, nearly fully expanded (mature) leaves requires all of the above plus diethyldithiocarbamate (DIECA) in the extraction medium. An extraction medium consisting of 25 mM EGTA, 0.01 M mercaptoethanol, 25 mM DIECA and 0.5% of the nonionic detergent, Nonidet-P40 (NP 40) was found to be very suitable for extraction of polysomes from all developmental stages of leaves. The polysomes extracted in the above medium showed active translation of protein in the wheat-germ in-vitro protein-synthesizing system. The translational products were similar when translations were carried out directly with polysomes or polysomal RNA, or polysomal poly(A)+ RNA from tobacco leaves. Poly(A)− polysomal RNA was a poor template in the in-vitro wheat-germ system.
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  • 28
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    Planta 152 (1981), S. 215-224 
    ISSN: 1432-2048
    Keywords: Bromodeoxyuridine tolerance ; Bromouracil excision ; Cytokinin autotrophy ; DNA synthesis ; Fluorodeoxyuridine effect ; Nicotiana
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract DNA isolated from various Nicotiana tabacum cell types, differing in their degree of hormone autotrophy and incubated in the presence of bromodeoxyuridine (BrdUrd), was analyzed by isopycnic CsCl gradient centrifugation. All cell types incorporate BrdUrd into DNA in such a way that hybrid DNA is formed with 60–80% of thymine (Thy) residues replaced by bromouracil (BrUra) in the newly synthesized strand. This DNA is not replicated further under ordinary culture conditions. Whereas in “normal” hormone-dependent cells this state is final and cells necrotize, in tumor (cytokinin-auxin autotrophic) and cytokinin-autotrophic cells a mechanism is induced leading to the reduction of BrUra content in DNA. As a result a decrease in the buoyant density (in CsCl) of BrUra DNA can be observed. In the case of cytokinin-autotrophic cells supplemented with kinetin, the buoyant density of the whole DNA decreases gradually to the value of that of unsubstituted DNA, but specific radioactivities of different DNA fractions reflect the retention of the pyrimidine ring of BrUra in DNA. This is interpreted as debromination of DNA in situ. The process can be inhibited by fluorodeoxyuridine (FdUrd) and deoxycytidine (dCyd). Moreover, FdUrd (but not dCyd) allows replication of hybrid DNA in tumor cells in such a way that HH DNA with all Thy residues replaced by BrUra is formed. For cytokinin-autotrophic cells FdUrd and kinetin are required. In hormone-dependent cells replication of hybrid DNA cannot be induced under any conditions. Most of these conclusions complement our previous findings that BrdUrd tolerance in hormone-autotrophic tobacco cells in hormone controlled. It is postulated that a modulation of thymidylate synthetase specificity is one factor affecting the level of BrUra substitution in DNA. The possibility of cytokinins being involved in the control of DNA synthesis is discussed.
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  • 29
    ISSN: 1432-2048
    Keywords: mRNA ; Nicotiana ; Protein synthesis ; Ribulose-1,5-bisphosphate carboxylase ; RNA (messenger) ; Translation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The products synthesized in vitro by messenger RNA (mRNA) extracted from Nicotiana sylvestris were analyzed by electrophoresis on polyacrylamide slab gels. Only three of the major polypeptides synthesized are considered here: P55, P32, and P20. P55 and P32 were translated from chloroplast mRNA. P55 corresponds to the large subunit of ribulose-1,5-bisphosphate (RuP2) carboxylase; P32 is probably a chloroplast membrane protein. P20, the polypeptide synthesized from cytoplasmic poly(A)+ RNA, is the precursor of the small subunit of RuP2 carboxylase. The balance between P20 and P32, in which their relative proportions varied inversely, was regulated by the age of the leaves and the time of illumination; we took advantage of this phenomenon to isolate the mRNA from the small subunit in relatively large amounts. This mRNA has a molecular weight of 350,000.
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  • 30
    ISSN: 1432-2048
    Keywords: Anther culture ; Fraction I protein ; Hybrid, somatic ; Nicotiana ; Ribulose-1,5-bisphosphate carboxylase
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    Topics: Biology
    Notes: Abstract Nine plants were produced from anthers of a somatic hybrid which had been obtained by fusion of Nicotiana tabacum L. and N. rustica L. protoplants. As determined by electrofocusing, the Fraction I protein of the original somatic hybrid had largesubunit polypeptides exclusively of the N. tabacum type. Two of the plants regenerated from anthers contained Fraction-I-protein large subunits exclusively of the N. rustica type. Since each plant was regenerated from a single cell, the somatic hybrid must have had cells containing both the N. tabacum and N. rustica chloroplast genome although the latter was not expressed. Possibilities to account for this non-expression of a chloroplast genome in the somatic hybrid are discussed.
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  • 31
    ISSN: 1432-2048
    Keywords: Adenosine 5′-phosphosulfate sulfotransferase ; Cell suspension culture ; Cysteine ; Enzyme regulation ; Nicotiana
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    Topics: Biology
    Notes: Abstract The properties and the regulation of adenosine 5′-phosphosulfate sulfotransferase extracted from cell suspension cultures ofNicotiana sylvestris was investigated. Optimal adenosine 5′-phosphosulfate sulfotransferase activity was obtained from the cells by extraction with 0.1 M tris-HCl, pH8.0, containing 2 M MgSO4 and 10 mM dithioerythritol. The K m for adenosine 5′-phosphosulfate in the sulfotransferase reaction was about 11 μM. Adenosine 5′-phosphosulfate in concentrations above 50 μM were inhibitory. The extratable adenosine 5′-phosphosulfate sulfotransferase activity decreased during cultivation with sulfate as the sole sulfur source, but after about 3 days it reached a constant level (50 to 100 nmol activated sulfate transferred h-1 mg-1 protein) which was maintained for at least 24 h. Addition of 0.5 mM cysteine to the culture medium decreased the extractable adenosine 5′-phosphosulfate sulfotransferase activity and blocked growth completely. With 0.1 mM cysteine an enzyme level of about 10% of the initial value was reached within 6 to 12 h without significant inhibition of growth. The added cysteine was absorbed rapidly and after 24 h cysteine could no longer be detected in the medium. Before the cysteine was completely depleted, the activity of adenosine 5′-phosphosulfate sulfotransferase started to increase, reaching ultimately a level which was comparable to the initial value.
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  • 32
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    Planta 147 (1980), S. 269-273 
    ISSN: 1432-2048
    Keywords: Crown gall ; Cytokinins ; Nicotiana
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    Topics: Biology
    Notes: Abstract Cytokinins were extracted from two cultures of tobacco crown gall tumor tissue: an unorganized tissue and a teratoma which produced leafy shoots. On Sephadex LH-20 column chromatography, extracts of both types of tissue yielded two peaks of cytokinin activity with elution volumes similar to ribosylzeatin and zeatin. Ribosylzeatin and zeatin were detected and quantified by coupled gas chromatography — mass spectrometry selected ion monitoring (GC/MS SIM), comparable quantities being found in the two extracts. Full mass spectral evidence for the presence of ribosylzeatin in both tissues was obtained. No evidence was found for the presence of N6-(Δ2-isopentenyl)adenosine (i6Ade) or N6-(Δ2-isopentenyl)adenine (i6Ade) although these compounds have been reported to occur in cytokinin-habituated tobacco callus tissues.
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  • 33
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    Planta 147 (1980), S. 274-276 
    ISSN: 1432-2048
    Keywords: Cell division ; Dikegulac ; Lysis ; Nicotiana ; Plasmalemma ; Protoplasts ; Tonoplast
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    Topics: Biology
    Notes: Abstract Dikegulac (2,3:4,6 di-o-isopropylidine-2-keto-I-gulonate) is a growth regulator used to differentially kill terminal apices, and it analogously inhibits basic metabolic functions in dividing cells, but not stationary cells, in suspension culture. This report demonstrates an analogous situation in isolated tobacco protoplasts. At the lowest concentrations, dikegulac partially suppresses division of the protoplasts. Higher concentrations are required to produce visual cytoplasmic damage to the protoplasts, which probably first occurs at the level of the plasmalemma, as the vacuoles can be released intact. Later, tonoplast disruption occurs.
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  • 34
    ISSN: 1432-2048
    Keywords: Lectin binding ; Membranes (surfaces) ; Nicotiana ; Organelles (leaf) ; Protoplasts ; Spinacia
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    Topics: Biology
    Notes: Abstract Organelles isolated from leaves of spinach (Spinacia oleracea L.) were prefixed in glutaraldehyde and then incubated with ferritin conjugates of four lectins — Concanavalin A (Con A), Ricinus communis L. agglutinin, MW 120,000 (RCA), soybean agglutinin (SBA), and wheat germ agglutinin (WGA) — in order to probe their cytoplasmic surfaces for saccharide residues. In each case the major leaf organelles, including microbodies, mitochondria and chloroplast derivatives, failed to exhibit labeling when examined with the electron microscope. Tobacco (Nicotiana tabacum L.) leaf protoplasts, incubated simultaneously with and under identical conditions to the spinach organelles, showed specific labeling of their plasma membranes with all four lectin conjugates, thus establishing the efficacy of the procedure for demonstrating the presence of binding sites when they exist. Further attempts to show binding of one of the lectins, Con A, by labeling with fluorescein-Con A and by organelle agglutination, yielded results consistent with the absence of ultrastructural labeling. It is concluded that no saccharide residues recognized by the four lectins are present on the cytoplasmic surfaces of organelles and that those residues reported to be constituents of intracellular membranes, therefore, are most likely exposed on the luminal (extracytoplasmic) surfaces.
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  • 35
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    Planta 148 (1980), S. 97-102 
    ISSN: 1432-2048
    Keywords: Abscisic acid ; Cell cultures ; Nicotiana ; Variants, resistant ; Water stress
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    Topics: Biology
    Notes: Abstract The goal of this work was to begin a genetic study of the molecular mode of action of abscisic acid (ABA), by isolating variant cultured cells resistant to the hormone, or to a factor which induces ABA synthesis, namely water stress. Cell cultures of Nicotiana tabacum L. cv. Wisconsin 38 and N. silvestris Speg. and Comes were chosen as the experimental materials. Studies of the effects of the two stresses on the growth of the cultures demonstrated that ABA or water stress imposed by mannitol could completely inhibit growth. These effects arose in both cases from a constant reduction of the growth rate of the cells throughout the culture period. Mannitol also induced an increase in ABA content of the cells and media of suspension cultures, although not to the concentrations required to achieve the same degree of growth inhibition when the hormone was applied exogenously.
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  • 36
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    Planta 148 (1980), S. 103-107 
    ISSN: 1432-2048
    Keywords: Abscisic acid ; Cell cultures ; Nicotiana ; Variants, resistant ; Water stress
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    Topics: Biology
    Notes: Abstract Variant clones were isolated from Nicotiana silvestris Speg. et Comes cell cultures at low frequencies following severe abscisic-acid (ABA) or mannitol-induced water-stress treatments of plated cells. N. tabacum L. variants were not recovered. Variants from the ABA selection experiments exhibited a 10-fold increase in resistance to the hormone. This trait was stable in non-selective conditions for as long as was tested (200 days), but did not alter the response of the cells to water stress. Cell lines from the waterstress selection were not more resistant to mannitol than the parent line, and had a wide range of response to ABA.
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  • 37
    ISSN: 1432-2048
    Keywords: mRNA ; Protein synthesis ; Protoplasts ; Nicotiana ; Translation
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    Topics: Biology
    Notes: Abstract Studies of proteins synthesized in vitro by messenger RNA (mRNA) extracted from tobacco protoplasts showed that the changes in protein synthesis and especially the lack of certain proteins observed previously in isolated protoplasts did not result from a failure of translation.
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  • 38
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    Planta 148 (1980), S. 462-467 
    ISSN: 1432-2048
    Keywords: Flower formation ; Leaves (in flower formation) ; Meristem (shoot) ; Nicotiana ; Roots (and flower formation)
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    Topics: Biology
    Notes: Abstract The terminal, apical shoot meristem ofN. tabacum cv. Wisconsin 38 normally differentiates into a flower after producing 30 to 40 nodes. The influence of leaves and roots on the regulation of flowering was evaluated by counting the number of nodes produced after removal of leaves or the induction of adventitious roots. Leaf removal has no effect on the number of nodes produced before flower formation. Root induction significantly increases the number of nodes produced before flower formation. The plant behaves as if it were measuring the number of nodes between the meristem and the roots as a means of regulating meristem conversion from vegetative to floral differentiation.
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  • 39
    ISSN: 1432-2048
    Keywords: Chloroplast ; Cybrid ; Nicotiana ; Protoplast ; Somatic hybrid
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    Notes: Abstract Protoplasts of a cytoplasmic albino mutant of Nicotiana tabacum L. characterized by a deficient chloroplast genome were fused with protoplasts of a nitrate-reductase deficient mutant (NR-) of N. tabacum. Somatic hybrids were obtained where the genome of the NR- mutant was complemented by the cytoplasmic albino mutant which could synthesize an active nitrate reductase, and the chlorophyll deficiency in the albino mutant was restored by the chloroplasts from the NR- mutant. Cybrids were also obtained in which the deficient plastids of the cytoplasmic albino mutant were replaced by normal chloroplasts from the NR- mutant. The system used permitted a simple selection of the hybrids and the cybrids. The NR- mutant was excluded at the cellular level by transfer of the cells to medium deficient in reduced nitrogen. The cytoplasmic albino mutant grew well on the selective nitrate medium. However, during callus formation, clear differences in the morphology and pigmentation of the calli were found which permitted selection for photoautotrophy at the callus level. The hybrid or cybrid nature of the plants was confirmed by examination of their morphology and chromosome number. Although the fusion partners come from the same species, only one plant showed the white-green variegated pattern typical of that of the cytoplasmic albino parent, indicating that segregation of plastids occurred during development of the calli and regeneration of the plants.
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  • 40
    ISSN: 1432-2048
    Keywords: 1-Aminocyclopropane-1-carboxylic acid ; Ethylene ; Light and ethylene production ; Nicotiana ; Pharbitis ; Protein synthesis
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    Topics: Biology
    Notes: Abstract Endogenous ethylene production of tobacco leaves was similar in light and in darkness. However, the rate of conversion of exogenously applied l-aminocyclopropane-l-carboxylic acid (ACC) to ethylene was reversibly inhibited by light. Virus-stimulated ethylene production, during the hypersensitive reaction of tobacco leaves to tobacco mosaic virus, was likewise inhibited by light. Under such circumstances ethylene production is limited at the level of the conversion of ACC to ethylene. Inhibition of the increase in ACC-stimulated ethylene production by cycloheximide and 2-(4-methyl-2,6-dinitroanilino)-N-methyl-propionamide after shifting leaf discs from light to darkness indicated that de novo protein synthsis was involved. Regulation of ACC-dependent ethylene production by reversible oxidation/reduction of essential SH groups, as suggested by Gepstein and Thimann (1980, Planta 149, 196–199) could be excluded. Instead, regulation of the ACC-converting enzyme at the level of both synthesis/degradation and activation/inactivation is suggested. Phytochrome was not involved in light inhibition, but low intensities of either red or blue light decreased the rate of ACC conversion. Dichlorophenyldimethylurea counteracted the inhibitory effect of light, indicating that (part of) the photosynthetic system is involved in the light inhibition. The ethylene production of Pharbitis cotyledons grown in darkness or light, either in the presence of absence of the inhibitor of carotenoid synthesis, SAN 9789 (norflurazon), supported this view.
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  • 41
    ISSN: 1432-2242
    Keywords: Nicotiana ; Somatic hybrids ; Protoplast fusion ; Genetic variability ; Variegation
    Source: Springer Online Journal Archives 1860-2000
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    Notes: Summary Somatic hybrids have been produced between N. tabacum and two closely related species in the genus Nicotiana, N. otophora and N. sylvestris, to evaluate interclonal variation and genetic behavior of these hybrids. As with the previously reported N. nesophila+ N. tabacum somatic hybrids, we have detected variation for morphological and isoenzyme characters between somatic hybrid clones, despite stability of chromosome number. One clone of N. sylvestris+N. tabacum was marked by variation in leaf spot frequency. The inheritance of this unstable trait was monitored through two sexual generations. Transmission of the Su gene marker was monitored in self-fertilized and back-crossed progeny of the N. sylvestris+N. tabacum somatic hybrids. Segregation ratios were similar to those previously reported for amphiploid N. sylvestris x N. tabacum sexual hybrids.
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  • 42
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    Theoretical and applied genetics 69 (1984), S. 1-14 
    ISSN: 1432-2242
    Keywords: Nicotiana ; Evolution of plastid DNA ; Comparative restriction site mapping ; Insertions and deletions
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Plastid DNA of seven American and four Australian species of the genus Nicotiana was examined by restriction endonuclease analysis using the enzymes Sal I, Bgl I, Pst I, Kpn I, Xho I, Pvu II and Eco RI. These endonucleases collectively distinguish more than 120 sites on N. tabacum plastid DNA. The DNAs of all ten species exhibited restriction patterns distinguishable from those of N. tabacum for at least one of the enzymes used. All distinctive sites were physically mapped taking advantage of the restriction cleavage site map available for plastid DNA from Nicotiana tabacum (Seyer et al. 1981). This map was extended for the restriction endonucleases Pst I and Kpn I. In spite of variation in detail, the overall fragment order was found to be the same for plastid DNA from the eleven Nicotiana species. Most of the DNA changes resulted from small insertions/deletions and, possibly, inversions. They are located within seven regions scattered along the plastid chromosome. The divergence pattern of the Nicotiana plastid chromosomes was strikingly similar to that found in the genus Oenothera subsection Euoenothera (Gordon et al. 1982). The possible role of replication as a factor in the evolution of divergence patterns is discussed. The restriction patterns of plastid DNA from species within a continent resembled each other with one exception in each instance. The American species N. repanda showed patterns similar to those of most Australian species, and those of the Australian species N. debneyi resembled those of most American species.
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  • 43
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    Theoretical and applied genetics 65 (1983), S. 269-276 
    ISSN: 1432-2242
    Keywords: Atropa ; Nicotiana ; cell hybrids ; Somatic hybrids ; Chromosome elimination ; DNA content
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    Topics: Biology
    Notes: Summary The genetic constitution of the cell hybrids Atropa belladonna + Nicotiana chinensis, obtained by cloning of individual heteroplasmic protoplast fusion products (Gleba et al. 1982) and cultured in vitro for 12 months, has been studied. The study comprised 11 hybrid cell clones of independent origin and included analysis of a) chromosome number, size, morphology, and relative position in metaphase plates, b) multiple molecular forms of the enzymes esterase and amylase, and c) relative nuclear DNA content. The data obtained permit us to conclude that, after one year of unorganized growth in vitro, the cells of most (8) clones had retained chromosomes of both parents, while species-specific elimination of nearly all Atropa chromosomes had occurred in three clones. About half of the non-segregating clones possess 120–150 chromosomes including 50–70 of Atropa and 50–90 of Nicotiana. Other clones are polyploid and possess 200–250 chromosomes with a predominance of either Atropa or Nicotiana chromosome types. Only a few chromosomal changes (reconstituted chromosomes, ring chromosomes) have been detected. In some metaphase plates, chromosomes of the two parents tend to group separately, indicating non-random arrangement of chromosomes of the two parents within the hybrid nucleus. Cytophotometric studies of the relative nuclear DNA content showed that distribution histograms for cell clones were similar to those of non-hybrid cultured cells. Cell populations were relatively homogenous and do not indicate any genetic instability as a result of hybridization between remote plant species. Biochemical analysis of isoenzyme patterns confirmed that in most cell clones, species-specific multiple molecular forms of esterase and amylase from both parents were present, i.e. genetic material of both parental species was expressed in the cell hybrids.
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  • 44
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    Theoretical and applied genetics 65 (1983), S. 213-217 
    ISSN: 1432-2242
    Keywords: Somatic hybridization ; Organelle segregation ; Nicotiana ; Male sterility ; Nitrate reductase ; Deficient mutant
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    Topics: Biology
    Notes: Summary Protoplasts from a nitrate reductase-deficient mutant of Nicotiana tabacum (cnx−68) were fused with protoplasts of 3 different cytoplasmically male-sterile cultivars of tobacco. Two cultivars had no stamens in the mature flowers and the third had petaloid structures in place of the stamens. Plants were regenerated from the fused protoplasts and characterized with respect to stamen development, chromosome number, and two chloroplast-coded traits. Nearly all hybrid plants displayed the chloroplast traits of only one parent, indicating that chloroplast segregation had occurred. The frequency of appearance of each chloroplast type differed according to the species origin of the chloroplasts. Chloroplasts from N. undulata competed much better than those from N. tabacum; N. suaveolens somewhat better than N. tabacum; and N. glauca about equally with N. tabacum. These results are compatible with an interpretation that equal frequency of appearance of chloroplast type among the regenerated plants occurs if the chloroplast DNAs of the parents are similar, whereas a bias of chloroplast type appears among the regenerated plants when the chloroplast DNAs are different. The appearance of aberrations in stamen development resembling the cytoplasmic male-sterile parental types was infrequent among the hybrid plants in all three crosses. Thus sterility factors were generally overcome by fertility factors following somatic hybridization.
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  • 45
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    Theoretical and applied genetics 64 (1982), S. 51-58 
    ISSN: 1432-2242
    Keywords: Haploid protoplast ; Diploid protoplasts ; Metabolic mutants ; U.V. sensitivity ; Nicotiana
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Protoplasts were derived from haploid and diploid Nicotiana sylvestris and N. tabacum. Exposure of the protoplasts to mutagenic doses of ultraviolet (U.V.) radiation prior to two selection rounds in the presence of 4 mM (or 5 mM) and 8 mM of valine, respectively, was required to obtain cell lines with persistent valine resistance. Such lines were obtained from haploid and diploid N. sylvestris protoplasts as well as from haploid protoplasts of N. tabacum but not from (1.8 × 107) diploid N. tabacum protoplasts. The ratio between number of verified valine-resistant cell lines and the initial number of U.V. exposed protoplasts enabled the estimation of the following order of mutation frequency: haploid N. sylvestris 〉 haploid N. tabacum 〉 diploid N. sylvestris. Plants which retained the valine resistance and transmitted it to their sexual progeny were derived from the resistant cell lines.
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  • 46
    ISSN: 1432-2242
    Keywords: Nicotiana ; Somatic hybrids ; Ribosomal RNA genes ; Restriction endonucleases
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    Notes: Summary Restriction endonucleases discriminated between the rDNAs contained in callus tissues derived from Nicotiana glauca, N. langsdorffii, and their somatic hybrids produced by protoplast fusion. With XbaI, a single repeat fragment of 7.5 × 106 daltons was produced from N. glauca rDNA compared to a single repeat fragment of 4.2 × 106 daltons produced from N. langsdorffii rDNA. Both kinds of XbaI fragments were found in somatic hybrids.
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  • 47
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    Theoretical and applied genetics 67 (1984), S. 491-497 
    ISSN: 1432-2242
    Keywords: Tobacco ; Chloroplast DNA ; Cybrids ; Male sterility ; Tentoxin resistance ; Plastid recombinants ; Nicotiana
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Protoplasts of a mutant line of Nicotiana tabacum having a maternally-transmitted chlorophyll deficiency were fused with protoplasts of two alloplasmic-male-sterile Nicotiana lines by the “donor-recipient” technique. In both fusion experiments variegated plantlets were regenerated which were shown to contain cytoplasms of mixed chloroplast nature. This confirms that with the “donor-recipient” method one can obtain mixed cytoplasms of genetically different chloroplasts. We present a convenient system to assay for genetic recombination between chloroplasts by combining use of several cytoplasmic markers: vis. chlorophyll pigmentation, chloroplast DNA restriction patterns, tentoxin resistance and male sterility. Within the limits of the experiment no recombinant types were recovered.
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  • 48
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    Theoretical and applied genetics 69 (1984), S. 121-128 
    ISSN: 1432-2242
    Keywords: Somatic cell ; Fusion ; Inheritance ; Nuclear genes ; Nicotiana
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Callus protoplasts of a Nicotiana tabacum chlorophyll-deficient mutant were fused with mesophyll protoplasts from one of following five sources: 4 cmsanalogs of tobacco bearing the cytoplasms of N. plumbaginifolia, N. suaveolens, N. repanda, and N. undulata, respectively, as well as wild species N. glauca. In another series of experiments, callus protoplasts from the chlorophyll-deficient genome Su/Su mutant of tobacco were fused with mesophyll protoplasts of the wild species N. glauca and those of a plastome chlorophyll-deficient tobacco mutant. The screening of hybrids consisted of visual identification followed by mechanical isolation and cloning of heteroplasmic fusion products in microdroplets of nutrient medium. Studies of regenerated plants included the analyses of gross morphology of plants, leaf and flower morphology, analysis of chromosome size and morphology and chromosome numbers, studies of multiple molecular forms of esterase and amylase, analysis of chloroplast DNA restriction patterns and analyses of chlorophyll-deficiency controlled by Su and P − genes. The study of progeny of 41 clones representing all species' combinations demonstrated that regenarants of most (63%) clones from intraspecific (for nuclear genes) combinations were cybrid forms, whereas in the case of the fusion N. tabacum + N. glauca, the true nuclear hybrids prevailed and the proportion of cybrids did not exceed 26%. Clones regenerating both hybrid and cybrid plants from the same fusion product were also found.
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  • 49
    ISSN: 1432-2242
    Keywords: Nicotiana ; Somatic hybrids ; Streptomycin resistance ; Plastid segregation ; Plastid DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Protoplasts of Nicotiana tabacum SRI (streptomycin resistant) and of Nicotiana knightiana (streptomycin sensitive) were fused using polyethylene glycol treatment. From three heterokaryons 500 clones were obtained. From the 43 which were further investigated, 6 resistant, 3 sensitive, and 34 chimeric (consisting of resistant and sensitive sectors) calli were found. From eight clones, a total of 39 plants were regenerated and identified as somatic hybrids. Chloroplast type (N. tabacum = NT or N. knightiana = NK) in the plants was determined on the basis of the species specific EcoRI restriction pattern of the chloroplast DNA. Regenerates contained NT (13 plants) or NK (15 plants) plastids but only the plants with the NT chloroplasts were resistant to streptomycin. This finding and our earlier data on uniparental inheritance points to the chloroplasts as the carriers of the streptomycin resistance factor.
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  • 50
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    Theoretical and applied genetics 60 (1981), S. 365-368 
    ISSN: 1432-2242
    Keywords: Disease resistance ; Isogenic lines ; Blackroot rot ; Thielaviopsis basicola ; Nicotiana
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Lines isogenic or near isogenic for traits other than resistance to black root rot from Nicotiana debneyi were developed in eight cultivar backgrounds of burley tobacco (N. tabacum L.). For each cultivar background, a resistant and susceptible selection from the seventh backcross generation plus the recurrent parental cultivar were evaluated for ten agronomic and chemical traits. Resistant selections were statistically different from the susceptible entries for days to flower, total nitrogen content, and total alkaloid content. Also, resistant selections were consistently lower in yield, but the differences were statistically nonsignificant. Resistant selections were also taller in three families and in two families the resistant selections had wider leaves. Linkage of genetic material from N. debneyi with the resistance factor was suggested as the possible reason for differences between resistant and susceptible selections.
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  • 51
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    Theoretical and applied genetics 61 (1982), S. 73-79 
    ISSN: 1432-2242
    Keywords: Nicotiana ; Chloroplast DNA ; Restriction fragments ; Deletion ; Evolution
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Nicotiana chloroplast genomes exhibit a high degree of diversity and a general similarity as revealed by restriction enzyme analysis. This property can be measured accurately by restriction enzymes which generate over 20 fragments. However, the restriction enzymes which generate a small number (about 10) of fragments are extremely useful not only in constructing the restriction maps but also in establishing the sequence of ct-DNA evolution. By using a single enzyme, Sma I, a elimination and sequential gain of its recognition sites during the course of ct-DNA evolution is clearly demonstrated. Thus, a sequence of ct-DNA evolution for many Nicotiana species is formulated. The observed changes are all clustered in one region to form a “hot spot” in the circular molecule of ct-DNA. The mechanisms involved for such alterations are mostly point mutations but inversion and deficiency are also indicated. Since there is a close correlation between the ct-DNA evolution and speciation in Nicotiana a high degree of cooperation and coordination betwen organellar and nuclear genomes is evident.
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  • 52
    ISSN: 1432-2242
    Keywords: Nicotiana ; Mitochondrial DNA-cybrids ; Somatic hybrids
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Mitochondrial DNA (mtDNA) restriction patterns were studied in mutant, cybrid and somatic hybrid plants regenerated from Nicotiana protoplasts. It has been shown that neither components of the culture media used for protoplast culture and plant regeneration, nor the antibiotics streptomycin and lincomycin used for the mutant selection induced alterations in the mtDNA. No rearrangements were detected in the mtDNA of plants derived from homoplasmic fusions where the mtDNA of the parents was identical as judged by mtDNA restriction patterns. There were rearrangements, however, in the mtDNA of each of the cybrid plants derived from heteroplasmic fusions. Restriction patterns generated by BamHI and SalI restriction endonucleases were different from those of both parents, and were composed of parental and non-parental fragments.
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  • 53
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    Theoretical and applied genetics 67 (1984), S. 185-193 
    ISSN: 1432-2242
    Keywords: Nicotiana ; Chloroplast DNA ; Deletion ; Restriction ; Hot spot ; Inverted repeats
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A physical map containing six restriction sites of the Nicotiana tabacum chloroplast genome, together with the BamHI maps of N. tabacum, N. otophora and N. knightiana, and the SmaI maps of N. acuminata, N. plumbaginifolia, N. langsdorffii, N. otophora, N. tabacum, N. tomentosiformis and N. knightiana was constructed. In Nicotiana chloroplast genomes, the most frequently observed variations are point mutations. Deletions are also detected. Most of the observed changes are confined to one area of the large single copy region, which is designated as the “hot spot”. Based on the evidence obtained from Nicotiana chloroplast genomes, an origin of the inverted repeats in this genus is proposed. We suggest that the inverted repeats represent a vestige of what were once two identical, complete chloroplast genomes joined together in a head-to-head and tail-to-tail fashion, and that deletions generated the current chloroplast genome organization.
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  • 54
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    Theoretical and applied genetics 67 (1984), S. 333-336 
    ISSN: 1432-2242
    Keywords: Nicotiana ; Chlamydomonas ; Chloroplast DNA ; RuBPCase ; E. coli ; B. subtilis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary RuBPCase, the enzyme responsible for carboxylation and oxidation of RuBP in a wide variety of photosynthetic organisms, is the major protein found in the chloroplast. Here we present the first evidence for direct expression in E. coli and B. subtilis of tobacco and Chlamydomonas ct-DNA sequences coding for the LS of RuBPCase as demonstrated by a simple in situ immunoassay.
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  • 55
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    Theoretical and applied genetics 58 (1980), S. 121-127 
    ISSN: 1432-2242
    Keywords: Nicotiana ; Cytoplasmic-male-sterility ; Somatic hybridization ; Plastid segregation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Restoration of male fertility was achieved by fusing protoplasts from male sterile (CMS) Nicotiana sylvestris plants with X-irradiated protoplasts derived from fertile N. tabacum plants. The CMS N. sylvestris plants were derived from a previous somatic hybridization experiment and contained alien (Line 92) cytoplasm. About one quarter of the regenerated plants were found to be cybrids. i.e. they consisted of N. sylvestris nuclei combined with all or some components of N. tabacum cytoplasm. In one half of these cybrids male fertility was restored to different levels. The chloroplasts of the two parental donors differ in respect to tentoxin sensitivity: chloroplasts of CMS N. sylvestris are sensitive while those of N. tabacum are insensitive. It could therefore be demonstrated that there was an independent segregation of chloroplast type and male fertility/sterility: several somatic cybrids were male fertile but tentoxin sensitive and others were tentoxin insensitive yet they were male sterile. Only in about one half of the somatic cybrids was male fertility restored together with restoration to tentoxin insensitivity.
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  • 56
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    Theoretical and applied genetics 62 (1982), S. 193-198 
    ISSN: 1432-2242
    Keywords: Nicotiana ; Somatic hybrids ; Protoplast fusion ; Ovule culture ; Genetic variability
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Somatic hybrids were produced between Nicotiana tabacum and N. nesophila, two species incapable of conventional sexual hybridization. Sexual hybrids, though, could be produced between these two species by using ovule culture only when N. nesophila was female. Clones of somatic hybrids were compared with sexual hybrids. Statistically significant variation was observed between clones, but not between sexual hybrids, for pollen viability, flower morphology, leaf morphology, and trichome density. As all clones of somatic hybrids have 96 chromosomes, the variability could not be explained by interclonal variation in chromosome number. Variation between somatic hybrids could be the result of cytoplasmic segregation or recombination, mitotic recombination or small chromosomal rearrangements prior to plant regeneration. Variation between clones could be exploited as these interspecies hybrids are now being used to incorporate disease resistance into cultivated tobacco.
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    Theoretical and applied genetics 63 (1982), S. 65-70 
    ISSN: 1432-2242
    Keywords: Disease screening ; Phytophthora parasitica var. ‘nicotianae’ ; Nicotiana ; Tissue culture ; Cytokinins
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Quantitatively inherited resistance to the black shank pathogen (Phytophthora parasitica var. ‘nicotianae’) was expressed among callus tissue cultures of tobacco (Nicotiana). Tissue cultures of genotypes known to posses polygenic mechanisms for black shank resistance expressed that resistance in vitro when challenged by the viable pathogen. Callus of a susceptible cultivar was readily parasitized in culture. Furthermore, single gene resistance to the common pathogen race was also shown to operate in vitro. Nongenetic factors examined did not contribute significantly to the observed differences. Disease expression in vitro appeared to be highly correlated with its expression at the whole plant level. Screening for quantitative disease resistance can be complicated at the whole plant level by variable hostpathogen reactions and by significant genotype × environment interactions. Since quantitatively inherited mechanisms of black shank resistance are expressed in tobacco callus cultures, an in vitro host-pathogen system may be useful in screening tobacco lines for black shank resistance.
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  • 58
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    Plant systematics and evolution 138 (1981), S. 89-113 
    ISSN: 1615-6110
    Keywords: Angiosperms ; Nicotiana ; Gossypium ; Lemnaceae ; Fraction 1 protein ; differentiation and age of taxa ; migration to Australia ; land connections ; long distance dispersal
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Fraction 1 protein (F-1-protein) (ribulose bisphosphate carboxy-lase-oxygenase) contained inLemnaceae has been evolving for at least 50 million years because fossils of these plants have been identified in strata belonging to the Upper Cretaceous. Electrofocusing F-1-protein resolves the large subunit polypeptides coded by extranuclear DNA and the small subunit polypeptides coded by nuclear DNA. Four differences affecting isoelectric points of the large subunit polypeptides and eight affecting the small subunit polypeptides are now present among eleven species representing the four genera comprising theLemnaceae. In comparison, four differences in the large and 13 in the small subunit polypeptides exist among 63 species ofNicotiana; four differences in the large and eight differences in the small subunit polypeptides exist among 19 species ofGossypium. The number of differences in F-1-protein composition being of the same order of magnitude for the generaNicotiana, Gossypium, and the familyLemnaceae, we infer that these Angiosperms are of similar antiquity. Nicotiana species indigenous to Australia and Africa contain F-1-proteins whose large subunit polypeptides are different but some of whose small subunit polypeptides are like those found in species from the Western Hemisphere. The same situation is found for the F-1-protein inGossypium. These results are in harmony with the view that species ofNicotiana andGossypium have arrived in Australia via former land connections between S. America, Antarctica, and Australia.
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  • 59
    ISSN: 1615-6102
    Keywords: Nicotiana ; Dark requirement ; Protoplast regeneration
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The protoplasts ofNicotiana plumbaginifolia required darkness for cell regeneration and colony formation. Maximal plating efficiency of the protoplasts could be achieved by keeping the cultures in dark instead of light or dark/light sequence. Only two days of darkness prior to the illumination at 400 or 3,000 lux resulted in appreciable plating efficiency, than those of light from the beginning, but these values could not match the high plating efficiency in total darkness.
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  • 60
    ISSN: 1615-6102
    Keywords: Cell cycle ; Membrane charge ; Nicotiana ; Plant protoplasts ; Tobacco mosaic virus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Attachment of virions of tobacco mosaic virus to protoplasts isolated from dividing suspension cultured cells ofNicotiana sylvestris was estimated using quantitative autoradiography of individual protoplasts. Additionally, the position of each protoplast in the cell cycle was assessed by Feulgen microspectrophotometry. At pH 5.6, after preincubation with 4 μg 1−1 poly-L-ornithine, protoplasts in the G1 and G2 phases bound more virions than protoplasts in the S-phase. The possibility that such differential binding was caused by cyclical variation in the net charge on the protoplast membrane has been investigated. It was found that S-phase protoplasts ofN. sylvestris can be separarated from protoplasts of other cycle stages by partition in aqueous, two-phase, immiscible polymer systems, presumably because they differ in charge. Also, electrophoretic studies suggest that G1 phase protoplasts bear higher surface charge than some non-G1 protoplasts.
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  • 61
    ISSN: 1615-6102
    Keywords: Nicotiana ; Somatic hybrids ; Chromosome ; Ribosomal ; RNA genes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The frequency of hybrid formation among calli developed from protoplasts ofNicotiana glauca andN. langsdorffii which had been treated with polyethylene glycole — Ca++ was determined. Approximately 5% of calli were proven to be somatic hybrids regardless selection pressure. Observation of chromosomes of selected hybrids showed variation in number ranging from 57–128, whereas parents contained 34–48. Analysis of ribosomal RNA genes (rDNA) confirmed every hybrid clone to contain rDNA from both parents.
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  • 62
    ISSN: 1615-6102
    Keywords: Embryogenic pollen ; Nicotiana ; Pollen culture ; Selection
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary By using density gradient centrifugation, employing 55% percoll and 4% sucrose as suspension medium, it is possible to select embryogenic pollen from buds after cold treatment at 10 °C for 8 or more days. These buds at the uninucleate stage of pollen were collected from plants grown in 8 hours photocycles at 18 °C and supplied with mineral salts. The embryogenic pollen are small, starch-free with a clear cytoplasm whereas large starch-filled ones are nonembryogenic. The embryogenic pollen regularly form embryos at a frequency of 2% on a mineral medium supplemented with glutamine, asparagine and sucrose at pH 6.5. These results demonstrate, for the first time, that it is possible to have embryos in appreciable frequencies in “ab initio” pollen cultures raised from cold treated anthers.
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  • 63
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    Plant cell, tissue and organ culture 2 (1983), S. 269-283 
    ISSN: 1573-5044
    Keywords: tissue culture ; protoplast fusion ; organogenesis ; isozyme analysis ; chloroplasts ; ribulose-1,5-bisphosphate carboxylase ; hybridization ; Nicotiana
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Protoplasts isolated from cell cultures of chlorophyll-deficient Nicotiana rustica cv. chlorotica and wild-type N. sylvestris were fused. The scheme for selection of somatic hybrids was based on the inability of the protoplast-derived colonies of the parental species to turn green; N. sylvestris protoplasts also had a very low plating efficiency in the medium employed. A total of 777 green colonies which were presumptive hybrids were isolated within four weeks of the fusion experiments. One hundred and eight green colonies formed shoots in vitro and 16 lines were rooted and grown in the greenhouse. Each of these hybrid plants displayed vegetative and floral traits intermediate to those of the parental species, except for plant height which in almost all cases was greater in the hybrids. Isozyme analyses by gel electrophoresis and isoelectric focussing of the small subunit of ribulose 1,5-bisphosphate carboxylase/oxygenase (RUBPCase) demonstrated that the nuclear genomes of both parents were expressed by the hybrids. Each of the eight somatic hybrid plants analyzed expressed only the N. rustica chloroplast genome as shown by isoelectric focussing of the large subunit of RUBPCase. This study demonstrated the value of N. rustica cv. chlorotica as a parental line in somatic hybridization with N. sylvestris and it might have widespread use with wild-type lines of other species.
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