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Showing papers on "Growth medium published in 1976"


Journal ArticleDOI
TL;DR: Evidence is presented that amino acid starvation is the specific stimulus initiating the developmental phase of the life cycle of Dictyostelium discoideum, and any effect of glucose on the primary control of the initiation of development is an indirect result of its utilization as a source of precursors for endogenously synthesized amino acids.

111 citations


Journal ArticleDOI
TL;DR: The yield of purified type III polysaccharide of group B Streptococcus was significantly improved by modification of the growth medium and a thicker microcapsule was found in organisms grown in standard broth.
Abstract: The yield of purified type III polysaccharide of group B Streptococcus was significantly improved by modification of the growth medium. Culture of organisms in standard Todd-Hewitt broth resulted in acid accumulation during the exponential phase of growth and poor yield of type III polysaccharide when extracted from cells by washing with neutral buffer solution. By increasing the buffering capacity of the broth medium, acid accumulation was prevented, and the number of viable cells was increased at the stationary phase of growth. Further, by increasing the concentration of glucose in the buffered medium, the yield of type III polysaccharide was increased two to three times. Electron microscopic investigations of cells grown in the modified broth medium demonstrated a thicker microcapsule than was found in organisms grown in standard broth.

70 citations


Journal ArticleDOI
TL;DR: It appears that the role of organic acids is neither connected to ammonium transport nor to relief of ammonia toxicity, but may be related to the need for additional carbon skeletons for synthesis of amino acids.
Abstract: Tobacco cells (Nicotiana tabacum) are capable of growth on ammonia as a sole nitrogen source only when succinate, malate, fumarate, citrate, alpha-ketoglutarate, glutamate, or pyruvate is added to the growth medium. A ratio between the molar concentrations of ammonia to succinate (as a complementary organic acid) in the growth medium of 1.5 was optimal. Succinate had no effect on the rate of uptake of ammonia from the medium into the cells although it did affect the intracellular concentration of ammonia. However, the changes were not sufficient to explain inhibition of growth as being due to ammonia toxicity. The radioactivity from (14)C-succinate was incorporated into malate, glutamate, and aspartate within 2 minutes.It appears that the role of organic acids is neither connected to ammonium transport nor to relief of ammonia toxicity, but may be related to the need for additional carbon skeletons for synthesis of amino acids.

52 citations


Journal ArticleDOI
TL;DR: Bean callus was induced to form roots (tissue differentiation) and vascular nodules (cell differentiation) by lowering the ratio of auxin to cytokinin in the growth medium and phenylalanine ammonia lyase activity of the calluses was closely correlated with the amount of cell differentiation which had occurred.
Abstract: Bean callus was induced to form roots (tissue differentiation) and vascular nodules (cell differentiation) by lowering the ratio of auxin to cytokinin in the growth medium. Both types of differentiation were inhibited by the addition of abscisic acid (at concentrations greater than I muM) to induction medium. Initiation of differentiation was inhibited, but its subsequent development was not, and the inhibition was not affected by the addition of gibberellic acid. Addition of gibberellic acid (GA) alone to induction medium stimulated tissue differentiation, although cell differentiation was unaffected (30 muM GA) or inhibited (45 muM GA) and its onset was delayed at both concentrations. Root initiation was also stimulated by gibberellic acid (0.I-45 muM) at an auxin-to-kinin ratio 10 times that normally optimal for cell differentiation. The phenylalanine ammonia lyase (PAL) activity of the calluses was closely correlated with the amount of cell differentiation which had occurred, and measurement of this confirmed that gibberellic acid delayed the initiation of cell differentiation. The increase and subsequent decline of PAL and betaI leads to 3 glucan synthetase activities, normally induced by transfer to induction medium, was abolished by abscisic acid. Addition of gibberellic acid did not affect the betaI leads to 3 glucan synthetase activity.

32 citations


Journal ArticleDOI
TL;DR: It was concluded that active amino acid transport in growing Streptomyces hydrogenans is under feedback control by intracellular amino acids.
Abstract: (1) The active uptake of different amino acids by growing cells of Streptomyces hydrogenans was shown to be correlated with the physiological age of the cells. During the lag phase of growth the transport capacity increased and attained its highest level when the growth rate was maximum. During further growth the transport capacity declined progressively. The lowest transport activity was observed when the culture shifted into the stationary growth phase. (2) Such modulation of transport capacity was independent on the presence or absence of amino acids in the growth medium of the cells. (3) The size and the composition of the pool of free intracellular amino acids was also undergoing substantial variations during the growth cycle of the culture. In the lag phase, the levels of all amino acids decreased markedly and attained their lowest values at the end of this phase. During further growth the pool size was slowly replenished. (4) Removal of the pool resulted in a considerable gain of transport capacity. Therefore, it was concluded that active amino acid transport in growing Streptomyces hydrogenans is under feedback control by intracellular amino acids. (5) Quantitatively, the modulation of the pool size could not fully account for the variation of the transport capacity. Since a pool-independent stimulation of transport was found to be correlated with the increase of the growth rate of the cells, the possibility is discussed that the stimulation of transport is either due to increased levels of distinct RNA species, which might provide positive feedback signals for transport, or by increased rates of de novo synthesis of transport limiting proteins.

29 citations


Journal ArticleDOI
TL;DR: It is suggested that the sugar composition of the growth medium has pronounced effects on cell physiology and that the effects are not mediated by cAMP.

28 citations


Journal ArticleDOI
TL;DR: 3-deoxyarabino-heptulosonic acid 7-phosphate synthase activity is derepressed in wild-type Escherichia coli K-12 cells grown on Fe3+-deficient medium; the phenylalanine-sensitive isoenzyme is not derepression under these conditions.
Abstract: 3-Deoxy-arabino-heptulosonic acid 7-phosphate synthase, prephenate dehydratase, tryptophan synthase, and 2,3-dihydroxybenzoylserine synthase enzyme activities are derepressed in wild-type Escherichia coli K-12 cells grown on Fe3+-deficient medium. This derepression is reversed when FeSO4 is added to the growth medium. Addition of shikimic acid to the Fe3+-deficient growth medium caused repression of the first three enzyme activities but not of 2,3-dihydroxybenzoylserine synthase activity. Addition of 2,3-dihydroxybenzoic acid to the Fe3+-deficient growth medium has no effect on any of the above-mentioned enzyme activities. The Fe3+ deficiency-mediated derepression of 3-deoxyarabino-heptulosonic acid 7-phosphate synthase activity is due to an elevation of the tyrosine-sensitive isoenzyme; the phenylalanine-sensitive isoenzyme is not derepressed under these conditions.

26 citations


Journal Article
TL;DR: The results are interpreted to suggest that melphalan transport by the L1210 leukemia cell is mediated by a system also responsible for the transport of glutamine and leucine and that interaction with such a system may play a significant role in the chemotherapeutic activity of this alkylating agent.
Abstract: Melphalan cytotoxicity to murine L1210 leukemia cells in culture was reduced in growth medium containing amino acids. Investigation of the effect of single amino acids revealed that the L-isomers of glutamine and leucine, but not the D-isomers, were the most active in decreasing cytotoxicity. Protection was concentration dependent, with maximum protection occurring at approximately 0.25 mM, a physiologic concentration. The LD90 for melphalan in the presence of 0.1 mM L-glutamine or L-leucine was increased by 7.3- and 10.8-fold respectively, under conditions where the cells had been pre-incubated with the amino acids. These results are interpreted to suggest that melphalan transport by the L1210 leukemia cell is mediated by a system also responsible for the transport of glutamine and leucine and that interaction with such a system may play a significant role in the chemotherapeutic activity of this alkylating agent.

24 citations


Journal ArticleDOI
TL;DR: It is concluded that Candida lipolytica, but not Saccharomyces cerevisiae, is able to adapt to the growth on fatty acids either by repression of fatty acid synthetase biosynthesis or by a fatty-acid-induced proteolytic degradation of the multienzyme complex.
Abstract: Endogeneous fatty acid biosynthesis in the two yeast species, Saccharomyces cerevisiae and Candida lipolytica is completely repressed by the addition of long-chain fatty acids to the growth medium. In Candida lipolytica, this repression is accompanied by a corresponding loss of fatty acid synthetase activity in the cell homogenate, when the cells were grown on fatty acids as the sole carbon source. The activity of the Saccharomyces cerevisiae fatty acid synthetase, however, remains unaffected by the addition of fatty acids to a glucose-containing growth medium. From fattyacid-grown Candida lipolytica cells no fatty acid synthetase complex can be isolated, nor is there any immunologically cross-reacting fatty acid synthetase protein detectable in the crude cell extract. From this it is concluded that Candida lipolytica, but not Saccharomyces cerevisiae, is able to adapt to the growth on fatty acids either by repression of fatty acid synthetase biosynthesis or by a fatty-acid-induced proteolytic degradation of the multienzyme complex. Similarly, the fatty acid synthetase complex disappears rapidly from stationary phase Candida lipolytica cells even after growth in fatty-acid-free medium. Finally, it was found that the fatty acid synthetase complexes from Saccharomyces cerevisiae and Candida lipolytica, though very similar in size and subunit composition, were immunologically different and had no common antigenic determinants.

23 citations


Journal ArticleDOI
TL;DR: The yield of parasynchronous cells is high with this technique and may produce a significant amount of nontemporally distorted biological material upon which direct biochemical analysis can be performed at various times within the generation cycle.
Abstract: A technique was investigated for producing parasynchronous growth of some established, aneuploid human cell strains. Removal of both serum and calcium from exponentially growing monolayer cells tended to inhibit their growth. After 20 hr, a high percentage of the cell population was arrested in or near mitosis. Readdition of serum and calcium caused parasynchronous growth of the cells of three human strains studied. All three strains incorporated tritiated thymidine maximally 10 to 15 hr after serum and calcium were added, and cell numbers increased rapidly 17 to 25 hr after the growth medium was reconstituted. Population-doubling ranged from 80% to 100% of the theoretical. The yield of parasynchronous cells is high with this technique and may produce a significant amount of nontemporally distorted biological material upon which direct biochemical analysis can be performed at various times within the generation cycle.

22 citations


Journal ArticleDOI
TL;DR: An unsaturated fatty acid-requiring mutant derived from Chinese hamster ovary (CHO) cells has been isolated and characterized and enzymatic defect of the mutant is localized to the microsomal stearoyl-CoA desaturase.
Abstract: An unsaturated fatty acid-requiring mutant derived from Chinese hamster ovary (CHO) cells has been isolated and characterized. This mutant grows normally when oleate or other unsaturated fatty acids are supplemented in the growth medium. Unlike the wild-type CHO cells, growth stops when medium is deprived of unsaturated fatty acid. Whole cell pulse experiments with [14C]acetate or [14C]stearate indicate that the mutant is defective in unsaturated fatty acid synthesis. Enzyme assays in vitro show that the enzymatic defect of the mutant is localized to the microsomal stearoyl-CoA desaturase.

Journal ArticleDOI
TL;DR: A germination medium was developed that suppressed microbial contamination and permitted long-term observation of these slow-germinating seeds and it was found that brief, intermittent light exposures depressed germination.
Abstract: A B S T RA CT Dormancy in seeds of the parasitic phanerogam Aeginetia indica L. can be broken by chemical treatment with sodium hypochlorite, which also helps to control contaminating microflora. A germination medium was developed that suppressed microbial contamination and permitted long-term observation of these slow-germinating seeds. The medium consisted of 10 ppm streptomycin, 10 ppm penicillin, and 10-' M indole-3-acetic acid (IAA) (or other growth regulator) in 1 % water agar. Optimum germination range was 25-30 C. Dormancy could also be broken by exposure on agar for several days at 3-5 C (stratification), or by brief exposures (15 min) to 50 C. Continuous light as low as 0.1 ft-c completely inhibited germination on this growth medium. Brief, intermittent light exposures depressed germination. Germination and growth in vitro of nondormant seed of Aeginetina indica L. can be described in five stages: (1) Germination: expansion of spheroidal cells or nodule at micropylar end of the seed, stimulated

Journal ArticleDOI
TL;DR: Sucrose agar is a relatively simple and inexpensive medium which gives good growth of a wide range of cultures of the Lactobacilli and Pediococci encountered in breweries and also of representative strains of other spoilage organisms.
Abstract: Sucrose agar is a relatively simple and inexpensive medium which gives good growth of a wide range of cultures of the Lactobacilli and Pediococci encountered in breweries and also of representative strains of other spoilage organisms. It may be used either for general purposes or, when modified to contain specific inhibitors, as a selective medium.

Journal ArticleDOI
TL;DR: It seems probable that mRNA synthesis is required for acquisition of increased transport activity and mRNA translation required for maintenance of normal activity and the controlling factor in the regulatory mechanism appears unlikely to be intracellular pool size.
Abstract: 1. Cultured cells were grown in various concentrations of amino acids for periods up to 3 days and the characteristics of the glycine transport system measured under fixed experimental conditions. During this time, the effect of enucleation, using cytochalasin B, and the effects of protein synthesis inhibitors (cycloheximide and actinomycin D) were investigated. 2. Glycine influx is regulated by the prior growth concentration of similarly transported amino acids. 3. The modification in transport involves primarily a change in Vmax (but also a change in Km in HeLa cells) and is effected within 2-10 hr after media change. Increased transport activity is calculated to be sufficient to compensate for the reduction in extracellular amino acid concentration, so that nearly normal influx values from media are maintained. Regulation over the range of concentrations studied is shown to be very accurate. 4. The nucleus is essential for the regulatory mechanism to function. It seems probable that mRNA synthesis is required for acquisition of increased transport activity and mRNA translation required for maintenance of normal activity. 5. The controlling factor in the regulatory mechanism appears unlikely to be intracellular pool size. Other possible signals are discussed.

Journal ArticleDOI
TL;DR: Radiorespirometric analyses revealed that vegetative cells of B. cereus metabolized glucose by simultaneous operation of the Embden-Meyerhof-Parnas pathway and the pentose phosphate pathway, and the extracellular products resulting from the metabolism of glucose decreased as the growth temperature was lowered.
Abstract: The influence of temperature on glucose metabolism of a psychotrophic strain of Bacillus cereus was investigated. The pH of the growth medium and spore-forming frequencies of B. cereus varied when grown at 32, 20, or 7 C. Radiorespirometric analyses revealed that vegetative cells of B. cereus metabolized glucose by simultaneous operation of the Embden-Meyerhof-Parnas pathway and the pentose phosphate pathway. As the growth temperature decreased, glucose was metabolized with increased participation of the pentose phosphate pathway. The shift of cells grown at a higher temperature to a lower temperature increased the relative participation of the pentose phosphate pathway, whereas the shift of cells grown at low temperatures to a higher temperature had the opposite effect. Cells of late logarithmic phase grown at 20 and 7 C oxidized acetate by the tricarboxylic acid cycle reaction. However, cells grown at 32 C failed to oxidize acetate to CO2 to any appreciable extent. The extracellular products resulting from the metabolism of glucose decreased as the growth temperature was lowered. Organic acids were the major extracellular products of cultures grown at 32 and 20 C. Acetic acid, lactic acid, and pyruvic acid together accounted for 86.1 and 78.9% of extracellular radioactivity, respectively, at the two temperatures. The relative ratio of these three acids varied between the temperatures. Little or no acid accumulated at 7 C.

Journal ArticleDOI
TL;DR: In buffered yeast extract L-asparagine medium, significant amounts of L-glutaminase and L- asparaginase activities appeared towards the end of the exponential phase and along the stationary phase and the process of enzyme formation showed a firm link to the cell active growth, as evidenced by the use of growth inhibitors.

Journal ArticleDOI
TL;DR: Experiments show that the amino acid pool can act as a nitrogen reserve but has little function as a carbon reserve, and at high NaCl concentrations there is a marked dependence for growth on the presence of sufficient potassium in the medium.
Abstract: The marine pseudomonad bacterium PL1 contains an intracellular pool of free amino acids which consist mainly of glutamate with small amounts of glutamine and aspartate when grown in a nutrient medium containing 0.2 M NaCl. When the NaCl concentration of the growth medium is increased to 0.8 M, proline becomes a major component of the intracellular pool together with glutamate—at this molarity and under suitable nutrient conditions these amino acids comprise 20% of total bacterial amino acid nitrogen. When grown in a nutrient growth medium containing a constant level of NaCl, the intracellular pool size can vary by a factor of 4 depending on the concentration of carbon and nitrogen in the medium. Experiments show that the amino acid pool can act as a nitrogen reserve but has little function as a carbon reserve. At high NaCl concentrations there is a marked dependence for growth on the presence of sufficient potassium in the medium. However, no correlation between K+ and glutamate concentration in either nitrogen or K+-limited cultures has been found. None of the enzymes associated with glutamate biosynthesis was influenced by NaCl levels between 0.2 and 0.5 M. Neither Na+ or K+ stimulated the activity of these enzymes when tested in vitro.

Journal ArticleDOI
TL;DR: The role of cyclic 3',Y-adenosine monophosphate (cAMP) has been well established in genetic control mechanisms and recently cAMP has been shown to increase the frequency of transformation in bacteria so it was of interest to investigate the role of exogenous cAMP on the growth of Neurospora.
Abstract: Also the paramorphogenic effect of cAMP was reversed by increasing the glucose concentration in the growth medium. Similar effect of glucose has been described in Escherichia coli [1]. When cAMP was added to the liquid culture, Neurospora wild-type strains were found to grow as small balls of mycelia. Thus the effect of cAMP on the growth pattern of Neurospora in liquid submerged culture was less pronounced. Effect of cAMP on conidiation: Besides its effect on the growth pattern of the wildtype strain cAMP and its dibutyryl derivatives were found to suppress the formation of aerial hyphae and conidiation in Neurospora. The wild-type strain grown on the minimal medium showed luxuriant condidiation after 30 h of growth whereas the culture grown on cAMP lacked aerial hyphae and conidiation even after 150h of growth. Effect o f cAMP on the sorbose-resistant mutant strains: It is known that the wildtype strain of Neurospora grows with colonial morphology in the presence of I.-sorbose [7]. However, a particular mutant strain called patch [8] is resistant to the paramorphogenic effect of L-sorbose. Furthermore it is known that sorbose can reduce the activity of wild-type cell-wall-synthesizing enzyme in vivo and in vitro but not that of the patch enzyme [9]. Therefore the effect of cAMP on the growth pattern The role of cyclic 3',Y-adenosine monophosphate (cAMP) has been well established in genetic control mechanisms [1]. Recently cAMP has been shown to increase the frequency of transformation in bacteria [2]. Since we have been interested in the problems o f genetic transformation in Neurospora [3, 4], it was therefore of interest to investigate the role of exogenous cAMP on the growth of Neurospora. The data presented in Table 1 show that cAMP induced a colonial-type growth in the wildtype strains of Neurospora (RL 3-8A RL21a). The wild-type strain showed a filamentous growth [5, 6] on the minimal medium containing glucose as sole carbon source. The growth pattern of the wildtype strain was changed from normal filamentous to semicolonial and finally to a colonial type when the increasing concentration of cAMP was added to the growth medium. The most dramatic effect was produced by cAMP (6 mg/ml) on solid agar medium. On such a medium, the wildtype strain showed a restricted colonial type of growth, similar effect was also seen when a dibutyryl derivative of cAMP was added to the growth medium. This paramorphogenic effect of cAMP and its dibutyryl derivative was characteristic of these nucleotides alone since other nucleotides such as 2' ,3 '-AMP, ADP and ATP were unable to affect the growth pattern of the Neurospora wild-type strain (see Table 1).

Journal ArticleDOI
TL;DR: It appears that the cell K is used to maintain cell [Na] below the NaCl concentration of the medium, and effects of low pH on cell ions are compared in metabolizing and starving bacteria, and it is shown that changes in the state of thecell K are correlated with movements of cell Na.
Abstract: Changes in cell volume and ion content of aHalobacterium species are described in terms of the NaCl concentration (0.5–3.5m) and pH (4–8) of the suspending medium. Cell volume, per unit content of protein of bacteria in stationary phase cultures, rose as the [NaCl] of the growth medium was increased. Logarithmic-phase bacteria shrank as the pH fell from 7 to 5.5. These changes are characteristic of bacteria with a moderate or rapid rate of O2 consumption. Starving (i.e. nonmetabolizing) bacteria, on the other hand, did not change in size within the above ranges of [NaCl] and pH. At lower values, however, such bacteria swelled and eventually lysed. Effects of low pH on cell ions are compared in metabolizing and starving bacteria, and it is shown that changes in the state of the cell K are correlated with movements of cell Na. It appears that the cell K is used to maintain cell [Na] below the NaCl concentration of the medium. The results are explained in terms of a model involving interactions between polyelectrolytes, salts and water in the concentrated cytoplasm of these halophilic organisms.

Patent
08 Apr 1976
TL;DR: In this paper, a process for biosynthetically producing cells in a liquid fermentation media comprised of a carbon source, and a growth medium containing oxygen and other essential cell nutrients, to obtain a biomass for harvesting is described.
Abstract: A process for biosynthetically producing cells in a liquid fermentation media comprised of a carbon source, and a growth medium containing oxygen and other essential cell nutrients, to obtain a biomass for harvesting. The carbon source and each of the other essential cell nutrients are added, incrementally or continuously, to the fermentation media, and each required nutrient is maintained at essentially the minimum level needed for efficient assimilation by the growing cells, in accordance with a predetermined cell growth curve based on the metabolic or respiratory function of the cells which convert the carbon source to a biomass. The process constitutes a marked improvement in accelerating and increasing cell production in a given fermentation system. In its preferred aspects, the nutrients are added within the fermentation media below the foam level which forms on top of the fermentation broth. A particularly preferred fermentation system embodies the preparation, in a nutrient medium, of comestible, digestible protein from a carbon source, suitably a hydrocarbon or carbohydrate, particularly delignified cellulose. A cellulase-elaborating microorganism, suitably a fungii, yeast or bacteria, particularly a microorganism of the genus Cellulomonas (ATCC-21399) used alone, or in compatible association with another microorganism, is cultivated within the medium which contains the carbon source.

Journal ArticleDOI
01 Nov 1976-Lipids
TL;DR: Fibroblast-like cells, derived from porcine aorta, were cultured under aerobic and anaerobic conditions and the fatty acid composition of the triacylglycerol content increased to 4 times the level determined in cells grown under aerobic conditions and remained constant throughout an additional 12 hr of growth.
Abstract: Fibroblast-like cells, derived from porcine aorta, were cultured under aerobic and anaerobic conditions. Light and electron microscopic examinations, lipid composition measurements, and incorporation of radioactive precursors into lipids of these cells were performed. Anaerobically grown cells accumulated oil red 0 stainable droplets and within 6 hr the triacylglycerol content increased to 4 times the level determined in cells grown under aerobic conditions. This ratio remained constant throughout an additional 12 hr of growth. The fatty acid composition of the triacylglycerols which accumulated under anaerobic conditions differed from the composition of fatty acids in the triacylglycerols present in the growth medium. The cellular unesterified fatty acids of the anaerobically grown cells differed only slightly in composition from the fatty acids in the growth medium, while the unesterified fatty acids of aerobically grown cells differed to a greater extent from those of the growth medium.

Journal ArticleDOI
TL;DR: If a trace amount of d-galactose-1-(14)C was supplied to cells after 15 days of growth in normal, high BA, or high 2,4-d media, there was no significant variation in uptake and utilization of label among the three growth conditions.
Abstract: Suspension cultures of Acer pseudoplatanus L. cells grown for 15 days in medium (T. Murashige and F. Skoog. 1962. Physiol. Plant. 15: 473-497) contained 3% sucrose, 1 mg/l 6-benzylaminopurine (BA), and 0.1 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D), referred to here as normal media, removed newly added myo-inositol-2-(3)H up to 100 mg/l in 24 hours and utilized up to 20% of this cyclitol for pectin biosynthesis. When the BA content of the growth medium was raised 10-fold, uptake of myo-inositol was drastically reduced and very little was available for pectin biosynthesis. Neither cell growth as measured by packed cell volume or by dry weight, nor monomer composition of pectic polysaccharides was affected by the increased level of cytokinin. Increasing, 2,4-d 10-fold instead of BA had little or no effect on myo-inositol uptake, although it did reduce the amount of myo-inositol utilized for pectin biosynthesis. Cells grown 15 days in normal media failed to remove added myo-inositol if 3% d-glucose was included. The net result was similar to that found in cells grown in the high BA condition. If a trace amount of d-galactose-1-(14)C was supplied to cells after 15 days of growth in normal, high BA, or high 2,4-d media, there was no significant variation in uptake and utilization of label among the three growth conditions.

Journal ArticleDOI
TL;DR: In medium from both untreated and X-irradiated cultures, a decrease in glutamic acid concentration was proportional to an increase in alanine concentration, and with time spent in culture, the amounts consumed and produced per cell were much higher in the irradiated cultures.
Abstract: KOVAL, T. M., MYSER, W. C., AND HINK, W. F. The Effect of X Irradiation on Amino Acid Utilization in Cultured Insect Cells. Radiat. Res. 67, 305-313 (1976). Cultured Trichoplusia ni cells in exponential growth were administered X-ray doses of 10,000 R and then subcultured. The concentration of alanine increased greatly and ammonia increased slightly in media of both untreated and irradiated cells up to 96 hr. Glycine concentration did not change significantly while the concentration of the remaining amino acids decreased to varying extents in media from both irradiated and untreated cultures. It was assumed that a decreasing amino acid concentration in the growth medium was due to utilization of the amino acids by the cells while an increasing amino acid concentration reflected its production by the cells. In medium from both untreated and X-irradiated cultures, a decrease in glutamic acid concentration was proportional to an increase in alanine concentration. With time spent in culture, the amounts consumed and produced per cell were much higher in the irradiated cultures.

Journal ArticleDOI
TL;DR: In the free-living nematode, Caenorhabditis briggsae, several enzymes involved in glycolysis change markedly in activity when the organism is incubated under different nutritional conditions, which can explain the differences in excretion products under both sets of conditions.
Abstract: 1. 1. In the free-living nematode, Caenorhabditis briggsae, several enzymes involved in glycolysis change markedly in activity when the organism is incubated under different nutritional conditions. 2. 2. Activities of phosphofructokinase and glycerol phosphate dehydrogenase increase sharply in worms incubated in “growth medium”. 3. 3. After incubation in buffer, the organisms show higher activities in aldolase, fructose-1,6-diphosphatase and glycerol phosphate dehydrogenase in the reverse direction (glycerol phosphate as substrate). 4. 4. These shifts in enzyme levels can explain the differences in excretion products under both sets of conditions. 5. 5. When C. briggsae is incubated with [2-14C]-acetate in buffer, glucose is the main radioactive product, but trehalose and small amounts of glycerol and fructose are also found; in whole medium, glycerol is most highly labeled, though detectable amounts of glucose and trehalose are present. 6. 6. Extracts of the worms yield similar products in both cases, namely, trehalose and glucose along with small amounts of ribose and glycerol.

Journal Article
TL;DR: A function of serum in the growth medium for primary monkey kidney cells has been shown to be inhibition of proteolytic enzymes, and a process of purifying and sterilizing trypsin without deleteriously affecting its proteolytically active activity is described.
Abstract: A function of serum in the growth medium for primary monkey kidney cells has been shown to be inhibition of proteolytic enzymes. Serum inactivates the residual trypsin remaining from enzymatic digestion of the kidneys and the proteolytic enzymes subsequently synthesized by the cells. Freshly trypsinized cells could be grown to monolayers in the absence of serum provided that they were repeatedly washed to remove residual trypsin. In the absence of serum, cell growth ceased on the 4-5th day after initiation of the culture, at which time the culture fluids became active proteolytically. When the 5th day fluids were replaced with fresh serum-free medium, cell growth was accelerated and a monolayer was attained by the 7th day. If cells were grown in the absence of whole serum but in the presence of medium containing alpha globulins or fetuin which inhibit both trypsin and cell proteases, such cultures grew as well as cultures containing serum. The sterilization of trypsin for use in digestion of tissues and cell cultures poses a serious problem. After filtration through 0.22 micron filters, trypsin preparations may still contain adventitious viruses, mycoplasma and minute forms of pseudomonas and other bacteria or bacteria-produced toxins, which pass the membrane pores. A process of purifying and sterilizing trypsin without deleteriously affecting its proteolytic activity is described.

Journal ArticleDOI
TL;DR: Clones at the 2 to 8 cell stage were mostly H-negative; this finding was consistent with the view that alterations in culture technique may have caused transient cellular and enzyme alterations which in the case of H antigen was reflected by a temporary loss of phenotype.

Journal ArticleDOI
TL;DR: Depletion of calf serum from the growth medium and addition of known quantities of lipids to the system provides a means of revealing subtle changes in lipid synthesis and lipid turnover during cellular growth.
Abstract: SummaryA simple medium system was developed to obtain growth of BHK-21 cells in shaker cultures in the absence of serum. These cells have now undergone over 80 serial passages in serum-free Way-mouth medium and have been recovered from the frozen state after storage for over 1 month in medium containing 10% dimethyl sulfoxide (DMSO) and 1% bovine serum albumin (BSA).Various amounts of exogenous lipid in the form of sodium oleate were added to cultures of cells growing in serum-free Way-mouth medium. Concentrations of 10–50 μg of sodium oleate/ml had no detrimental effects on the cells as measured by trypan blue uptake. Furthermore, the cells were serially passed ten times in the presence of 10 μg sodium oleate/ml. Depletion of calf serum from the growth medium and addition of known quantities of lipids to the system provides a means of revealing subtle changes in lipid synthesis and lipid turnover during cellular growth.The technical assistance of K. Crilly is gratefully acknowledged.

Journal ArticleDOI
TL;DR: A strong correlation between exogenously supplied adenine and related compunds, increased growth, increased protein kinase activity, and kinase response to cAMP in this organism is demonstrated.
Abstract: Growth ofVerticillium albo-atrum in liquid Czapek-Dox broth was stimulated about four-fold by added 10 mM adenine, N6-benzyladenine, or kinetin. Less stimulation was evident at lower concentrations. With none of these included in the basal growth medium, detectable protein kinase activity in cell-free extracts was low and responded minimally to cAMP (adenosine 3′, 5′-cyclic monophosphate) in the reaction mixture. With each of these compounds as an additive to the growth medium, protein kinase activity was not only greater but also responded markedly (several fold) to cAMP. These results demonstrate a strong correlation between exogenously supplied adenine and related compunds, increased growth, increased protein kinase activity, and kinase response to cAMP in this organism.

Journal ArticleDOI
TL;DR: Time-course studies showed that there is no correlation between the cellular concentrations of either polyphosphate or orthophosphate and the ability of the cells to form this enzyme.
Abstract: The relationship between the level of stored polyphosphate in growing cells of Saccharomyces bisporus and the repression or derepression of the synthesis of the enzyme acid phosphatase (EC 3.1.3.2) was investigated. Time-course studies showed that there is no correlation between the cellular concentrations of either polyphosphate or orthophosphate and the ability of the cells to form this enzyme. The only compound investigated that was capable of repressing acid phosphatase synthesis was orthophosphate in the growth medium (i.e., orthophosphate outside the cell).

Journal ArticleDOI
TL;DR: The finding that pyruvate added to minimal growth medium causes cellular flattening and formation of external processes accompanied by increaded DNA synthesis in cultured hepatoma cells (HTC cells) suggests that alterations in energy metabolism, or precursors that enhance viability and macromolecular synthesis in mammalian cell cultures, may exert marked effects on cellular morphology without corresponding changes in growth of neoplastic liver cells.
Abstract: Cells maintained in basal growth medium with 02–10% serum often require citric acid cycle intermediates for optimal viability We have found that pyruvate added to minimal growth medium causes cellular flattening and formation of external processes accompanied by increaded DNA synthesis in cultured hepatoma cells (HTC cells) Cells were cultured in plalstic T-flasks (05, 10, or 20 × 106 cells/flask) containing 5 ml medium (90% Eagle's Basal Medium (BME) and 10% Swim's S-77) with various concentrations of fetal calf serum (02,025, 05, 10, 20, 10%) and either pyruvate (50, 100, 250,500, 1,000μg/ml), or one of: dibutyryl cAMP (DBcAMP) or dibutyryl cGMP (DBcGMP) at 10−3, 10−4, or 10−5 M At 44–48 hr cultures were pulsed with tritiated thymidine, uridine, or lecucine Cells became attached to the plastic surface within 24hr Cells in medium with 025 to 20% serum had a rounded appearance With added pyruvate, cellular flattening, process formation, and an increased adherence to the substratum was absorbed By 48 hr, culture without pyruvate grew in rounded clusters; with pyruvate, cells formed extensive interconnecting processes that appeared loosely attached to the monolayer surface At the cell densities tested, process formation was maximal with 250 to 500 μg/ml pyruvate Cytochalasin B blocked flattening and process formation; EDTA (1 mg/ml) caused retraction of processes within 3 min, and a slow dissolution of these structures within cells was observed DBcAMP or DBcGMP did not induce process formation Flattening and process foormation in pyruvate-enriched cultures were accompanied by marked stimulation of DNA synthesis and smaller increases in RNA and protein synthesis Cell number was not affected These pyruvate-induced changes suggest that alterations in energy metabolism, or precursors that enhance viability and macromolecular synthesis in mammalian cell cultures, may exert marked effects on cellular morphology without corresponding changes in growth of neoplastic liver cells