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41.
Cost estimates have been prepared for commercial-scale production of ajmalicine-rich Catnaranthus roseus biomass using plant cell culture. At the current state of the technology the cost would be approximately $7.30/lb dry biomass ($3215/kg ajmalicine). Naturally-grown C. roseus roots have a 50% lower ajmalicine concentration but would cost only ca. $0.70/lb ($619/kg ajmalicine). The principal reason for the high cost of the plant cell culture route is not the slow specific growth rate (0.35 day(-1)), but rather the slow specific product accumulation rate (0.26 mg/g day). This rate will have to be increased by a factor of 40 to make the process competitive. 相似文献
42.
A model for growth of Saccharomyces cerevisiae containing a recombinant plasmid in selective media 总被引:1,自引:0,他引:1
A major problem in the use of plasmids as recombinant vectors is the problem of plasmid-free cell generation from plasmid shedding and subsequent growth. A common technique for controlling the population of plasmidfree cells is the use of selective media against these cells using an auxotrophic host and a plasmid that has the ability to produced the essential metabolite. A distributed model describing the growth of Saccharomyces cerevisiae containing a recombinant plasmid in selective media was developed. The model allows for growth and production of a metabolite by the plasmid-carrying strain and growth of the plasmid-free cells on resulting metabolite concentrations. Through a determination of system constants and numerical solution to the equations, experimental batch and continuous culture results for cell concentration transients could be simulated by the model. The results indicated that despite selective pressure, plasmid-free cell growth was significant. 相似文献
43.
Photosynthetic ATP regeneration was measured in open reactors using immobilized Chromatophores from Rhodopseudomonas capsulata. The influence of several factors on both initial and long-term ADP photophosphorylation was studied. The effect of phosphate salts and of bovine serum albumin on the organelle activity yield was studied. Photophosphorylation was initiated either with ADP or regenerated ATP and the roles of these nucleotides were compared. Different photoreactor configurations were tested for the production of a phosphorylated compound and a flat reactor selected. The presence of inorganic pyrophosphate in the reaction medium was shown to improve the synthesis of ATP 1.4 times. Using the optimal conditions described here, the total G-6P production was 50-fold higher than in batch reactors. 相似文献
44.
Symplastic Transport of Carboxyfluorescein in Staminal Hairs of Setcreasea purpurea Is Diffusive and Includes Loss to the Vacuole 总被引:8,自引:4,他引:4
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The kinetics of symplastic transport in staminal hairs of Setcreasea purpurea was studied. The tip cell of a staminal hair was microinjected with carboxyfluorescein (CF) and the symplastic transport of this CF was videotaped and the digital data analyzed to produce kinetic curves. Using a finite difference equation for diffusion between cells and for loss of dye into the vacuole, kinetic curves were calculated and fitted to the observed data. These curves were matched with data from actual microinjection experiments by adjusting K (the coefficient of intercellular junction diffusion) and L (the coefficient of intracellular loss) until a minimum in the least squares difference between the curves was obtained. (a) Symplastic transport of CF was governed by diffusion through intercellular pores (plasmodesmata) and intracellular loss. Diffusion within the cell cytoplasm was never limiting. (b) Each cell and its plasmodesmata must be considered as its own diffusion system. Therefore, a diffusion coefficient cannot be calculated for an entire chain of cells. (c) The movement through plasmodesmata in either direction was the same since the data are fit by a diffusion equation. (d) Diffusion through the intercellular pores was estimated to be slower than diffusion through similar pores filled with water. 相似文献
45.
Molecular Cloning of Osmotin and Regulation of Its Expression by ABA and Adaptation to Low Water Potential 总被引:40,自引:12,他引:28
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In response to adaptation to NaCl, cultured tobacco cells (Nicotiana tabacum L. cv Wisconsin 38) synthesize a major 26 kilodalton protein which has been named osmotin due to its induction by low water potentials. To help characterize the expression of osmotin in adapted cells, a cDNA clone for osmotin has been isolated. Abscisic acid induces messenger RNA encoding osmotin. Levels of this mRNA in adapted cells are approximately 15-fold higher than in unadapted cells. Message for osmotin is present at constant levels through the growth cycle of adapted cells, while in unadapted cells, the level decreases during exponential phase of growth and increases again when the cells approach stationary phase. While abscisic acid induces the message for osmotin, a low water potential environment appears to be required for accumulation of the protein. An osmotic shock to unadapted cells does not increase the amount of message or protein present most likely because this treatment does not induce immediately the accumulation of abscisic acid. The increased expression of osmotin in adapted cells is not correlated with an increase in osmotin gene copy number. Osmotin is homologous to a 24 kilodalton NaCl-induced protein in tomato, as well as thaumatin, maize α-amylase/trypsin inhibitor and a tobacco mosaic virus-induced pathogenesis-related protein. 相似文献
46.
Characterization of the Effects of Divalent Cations on the Coupled Activities of the H-ATPase in Tonoplast Vesicles 总被引:4,自引:2,他引:2
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The substrate requirement of the H+-ATPase in purified corn root tonoplast vesicles was investigated. The coupled activities, ATP hydrolysis and proton pumping, were simultaneously supported only by Mg2+ or Mn2+. The presence of Ca2+ or Ba2+ did not significantly affect the coupled activities. The addition of Cd2+, Co2+, Cu2+, and Zn2+ inhibited both the hydrolysis of Mg-ATP and the proton transport. However, the inhibition of proton pumping was more pronounced. Based on equilibrium analysis, both ATP-complexed and free forms of these cations were inhibitory. Inhibition of the hydrolysis of Mg-ATP could be correlated to the concentrations of the ATP-complex of Zn. On the other hand, the free Cu2+ and Co2+ were effective in inhibiting hydrolysis. For proton pumping, the ATP complexes of Co2+, Cu2+, and Zn2+ were effective inhibitors. However, this inhibition could be further modulated by free Co2+, Cu2+, and Zn2+. While the equilibrium concentrations of Cd-ATP and free Cd2+ were not estimated, the total concentration of this cation needed to inhibit the coupled activities of the H+-ATPase was found to be in the range of 10 to 100 micromolars. The presence of free divalent cations also affected the structure of the lipid phase in tonoplast membrane as demonstrated by the changes of emission intensity and polarization of incorporated 1,6-diphenyl-1,3,5-hexatriene. The differential inhibition caused by these cations could be interpreted by interactions with the protogenic domain of the membrane as previously proposed in “indirect-link” mechanism. 相似文献
47.
Hydroxycinnamoyl-CoA:Putrescine Hydroxycinnamoyltransferase in Tobacco Cell Cultures with High and Low Levels of Caffeoylputrescine 总被引:1,自引:0,他引:1
A new hydroxycinnamoyl-CoA:putrescine hydroxycinnamoyltransferase (PHT) was detected in two variant lines of Nicotiana tabacum L. (TX1, TX4) accumulating markedly different levels of caffeoylputrescine. The enzyme accepted only the aliphatic diamines putrescine, cadaverine and 1,3-diaminopropane at a ratio of 100:33:8. Caffeoyl- and feruloyl-CoAs were the best acyl donors. The apparent Km-values for caffeoyl-CoA and putrescine were near 3 and 10 micromolar, respectively, at the pH-optimum of 10.0. PHT activity was quite similar in low producing TX1 and high producing TX4 cells, while some other biosynthetic enzymes (phenylalanine ammonia-lyase, ornithine decarboxylase) were greatly enhanced in TX4 cells, suggesting that PHT does not catalyze the rate-limiting step in hydroxycinnamoylputrescine formation. 相似文献
48.
Evidence for a ferredoxin-dependent choline monooxygenase from spinach chloroplast stroma 总被引:18,自引:5,他引:13
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Chenopods synthesize betaine in the chloroplast via a two-step oxidation of choline: choline → betaine aldehyde → betaine. Our previous experiments with intact chloroplasts, and in vivo18O2 labeling studies, led us to propose that the first step is mediated by a monooxygenase which uses photosynthetically generated reducing power (C Lerma, AD Hanson, D Rhodes [1988] Plant Physiol 88: 695-702). Here, we report the detection of such an activity in vitro. In the presence of O2 and reduced ferredoxin, the stromal fraction from spinach (Spinacia oleracea) chloroplasts converted choline to betaine aldehyde at rates similar to those in intact chloroplasts (20-50 nanomoles per hour per milligram protein). Incorporation of 18O from 18O2 by the in vitro reaction was demonstrated by fast atom bombardment mass spectrometry. Ferredoxin could be reduced either with thylakoids in the light, or with NADPH plus ferredoxin-NADP reductase in darkness; NADPH alone could not substitute for ferredoxin. No choline-oxidizing activity was detected in the stromal fraction of pea (Pisum sativum L.), a species that does not accumulate betaine. The spinach choline-oxidizing enzyme was stimulated by 10 millimolar Mg2+, had a pH optimum close to 8, and was insensitive to carbon monoxide. The specific activity was increased threefold in plants growing in 200 millimolar NaCl. Gel filtration experiments gave a molecular weight of 98 kilodaltons for the choline-oxidizing enzyme, and provided no evidence for other electron carriers which might mediate the reduction of the 98-kilodalton enzyme by ferredoxin. 相似文献
49.
Transformation of Brassica napus and Brassica oleracea Using Agrobacterium tumefaciens and the Expression of the bar and neo Genes in the Transgenic Plants 总被引:25,自引:3,他引:22
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An efficient and largely genotype-independent transformation method for Brassica napus and Brassica oleracea was established based on neo or bar as selectable marker genes. Hypocotyl explants of Brassica napus and Brassica oleracea cultivars were infected with Agrobacterium strains containing chimeric neo and bar genes. The use of AgNO3 was a prerequisite for efficient shoot regeneration under selective conditions. Vitrification was avoided by decreasing the water potential of the medium, by decreasing the relative humidity in the tissue culture vessel, and by lowering the cytokinin concentration. In this way, rooted transformed shoots were obtained with a 30% efficiency in 9 to 12 weeks. Southern blottings and genetic analysis of S1-progeny showed that the transformants contained on average between one and three copies of the chimeric genes. A wide range of expression levels of the chimeric genes was observed among independent transformants. Up to 25% of the transformants showed no detectable phosphinotricin acetyltransferase or neomycin phosphotransferase II enzyme activities although Southern blottings demonstrated that these plants were indeed transformed. 相似文献
50.
Procedures for the Generation of Mature Chlamydomonas reinhardtii Zygotes for Molecular and Biochemical Analyses
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Zygotes represent an important stage in the sexual cycle of the unicellular green alga Chlamydomonas reinhardtii. To study zygote germination at a molecular level, a protocol was elaborated for the generation of zygotes in large quantities and a method was developed for the extraction from zygotes of RNA that could be translated in vitro. 相似文献