Showing papers on "Growth medium published in 1998"

Extracellular products as mediators of the formation and detachment of Pseudomonas fluorescens biofilms

Abstract: Pseudomonas fluorescens B52 produces substantial biofilms at the air/liquid/solid interface of glass coverslips clamped vertically and partly submerged in liquid medium at 21 degrees C. Biofilm formation was maximal ca. 20-50 h after inoculation of the liquid medium and as indicated by environmental scanning electron microscopy (ESEM), contained large numbers of bacterial cells that were embedded within an extensive exopolymeric matrix. Incubation beyond 50 h led to reductions in biofilm which ESEM related primarily to losses of exopolymer. Both biofilm formation and the subsequent decline in exopolymer deposition was more rapid, and occurred to greater extents, when supernatants from two-day old cultures of B52 were used as the initial growth media. The addition of N-acyl-hexanoyl homoserine lactone to fresh growth medium had a similar effect upon biofilm formation as using spent culture medium. Homoserine lactones could not be demonstrated in spent culture supernatants by an Agrobacterium tumefaciens bioassay. An exopolysaccharide lyase was detected in spend culture media taken from dense biofilm cultures whose action was specifically directed towards biofilm exopolysaccharide. Results suggest that (i) cell-cell signals such as homoserine lactones are associated with the formation of P. fluorescens biofilms, (ii) the enzymic degradation of exopolymers has a specific role in the detachment of cells under starvation conditions, and (iii) whilst short chain (C6) exogenous homoserines can trigger such response in P. fluorescens, its own signal substance is likely to possess a longer (> C8) fatty acyl chain.

Elemental balancing of biomass and medium composition enhances growth capacity in high-density Chlorella vulgaris cultures.

Abstract: The basic requirements for high-density photoautotrophic microalgal cultures in enclosed photobioreactors are a powerful light source and proper distribution of light, efficient gas exchange, and suitable medium composition. This article introduces the concept of balancing the elemental composition of growth medium with biomass composition to obtain high-density cultures. N-8 medium, commonly used for culturing Chlorella vulgaris was evaluated for its capacity to support high-density cultures on the basis of elemental stoichiometric composition of C. vulgaris. This analysis showed that the N-8 medium is deficient in iron, magnesium, sulfur, and nitrogen at high cell densities. N-8 medium was redesigned to contain stoichiometrically balanced quantities of the four deficient elements to support a biomass concentration of 2% (v/v). The redesigned medium, called M-8 medium, resulted in up to three- to fivefold increase in total chlorophyll content per volume of culture as compared to N-8 medium. Further experiments showed that addition of each of the four elements separately to N-8 medium did not improve culture performance and that balanced supplementation of all four deficient elements was required to yield the improved performance. Long-term (24 d) C. vulgaris culture in M-8 medium showed continuous increase in chlorophyll content and biomass throughout the period of cultivation. In contrast, the increase in chlorophyll content and biomass ceased after 7 and 12 d, respectively in N-8 medium, demonstrating the higher capacity of M-8 medium to produce biomass. Thus, the performance of high cell density photobioreactors can be significantly enhanced by proper medium design. The elemental composition of the biomass generated is an appropriate basis for medium design.

Stress tolerance in the marsh plant Spartina patens: Impact of NaCl on growth and root plasma membrane lipid composition

Abstract: The C-4 salt marsh grass, Spartina patens, thrives in the upper portion of the marsh where soil salinities may be equal to coastal seawater. Spartina patens was grown in hydroponic culture in a greenhouse at 0, 340, and 510 mM NaCl, and measured for growth, tissue cation content, and root plasma membrane (PM) lipid composition. From 0 to 340 and 510 mM, the shoot growth decreased, but root growth was not affected. The Na + content increased in both shoots and roots when plants were grown in salt, while the shoots had a decreased K + content and the roots had a decreased Ca 2+ content. Spartina patens root plasma membrane was isolated with an aqueous polymer two-phase system. The purity of the plasma membrane was verified with cytochemical tests on membrane enzyme markers. Plasma membrane lipids were stable relative to the membrane protein content. Molar percentages of sterols (including free sterols) and phospholipid decreased with increasing salinity. However, glycolipid showed a statistically significant increase in the total lipid as salinity in the medium was increased from 0 to 510 mM. Even at a salinity of 510 mM, the plasma membrane sterol/phospholipid ratio was unaffected by NaCI. When the plants were grown in NaCI media, the plasma membrane had a decreased phosphatidylcholine (PC) and phosphatidylethanolamine (PE) content, but the PC/PE ratios were not affected. The plasma membrane molar percentage of sitosterol in total free sterol increased when plants were grown in salt media. The predominant membrane fatty acids were C11 and C14, and the major unsaturated one was C14:1. An increase in growth medium salinity resulted in a decreased root plasma membrane fluidity.

On the iron requirement of lactobacilli grown in chemically defined medium

Abstract: The iron requirement of four strains of lactobacilli (L. acidophilus, L. delbrueckii subsp. bulgaricus, L. plantarum, and L. pentosus) was studied in a synthetic medium under aerobic or anaerobic conditions. Effects of iron salt and iron-chelated compounds were tested on bacterial growth in manganese-free or -supplemented media. No significant growth stimulation was observed in any condition. These results support the absolute manganese requirement for optimum growth of lactobacilli and the needless incorporation of iron in growth media.

Single cell protein from pineapple cannery effluent

Abstract: Candida utilis was grown on pineapple cannery effluent as the sole carbon and energy source. These effluents are rich in microbially utilizable nutrients. The principal sugars in the effluent were glucose, fructose and sucrose, and when supplemented with diammonium hydrogen phosphate to prevent nitrogen limitation they supported good growth. The maximum specific growth rate (μm=0.46h–1) and cell yield coefficient (Yc/s=0.30) were obtained with 23.2 g carbohydrate/l in the growth medium. The values of μm and Yc/s varied with carbohydrate concentration: higher values of μm and Yc/s were obtained with lower concentrations of carbohydrate, suggesting that a chemostat would be more suitable for single cell protein production. Freeze-dried yeast contained 55.3, 51.2, 1.45, 6.4 and 27.4% of crude protein, true protein, DNA, RNA and carbohydrate respectively. The yeast had a balanced amino acid profile, except for sulphur-containing amino acids. A 90–95% reduction in the chemical oxygen demand of the effluent was achieved during this process which suggests that yeast may be effectively used to treat this waste.

Optimization of the mineral composition of in vitro culture media

Abstract: Plant tissue culture requires the optimization of growth media. The effect of eight different macronutrient formulations on the in vitro culture of Gatharanthus roseus nodes was investigated: the caulogenesis rate decreased with the overall ion concentration and the ammonium content of the growth medium. The methodology proposed for the optimization of the mineral composition of the growth medium is based upon the adjustment of the macronutrients contents to the plant requirements. Such an approach was used for the comparison between the effects of the optimized growth medium and those of the usual Murashige and Skoog solution in the growth of the multiple shoots issued from Solatium paludosum root calluses. The new formulation MH1 [with a diminution of the ionic concentration, nitrogen (N) supplied in the form of nitrate and calcium (Ca) in larger amounts than potassium (K)] doubled the biomass production and tripled the yield of solamargine, a secondary metabolite.

Cyanophycin and glycogen synthesis in a cyanobacterial Scytonema species in response to salt stress

Abstract: The response to salinity of a Scytonema species isolated from the central Australian desert was studied. Under nitrogen-fixing conditions the addition of increasing concentrations of salt (NaCl) caused progressive inhibition of growth, with growth ceasing at 150 mM NaCl. This correlated with a progressive loss of nitrogenase activity, a low level of activity being retained at 150 mM NaCl. The inhibition of growth was overcome when KNO3 (10 mM) was added to the growth medium. In response to the salt stress, cells accumulated the reserve compounds cyanophycin and glycogen. Time course experiments showed that they were steadily synthesized over 48 h, after which the concentrations stabilized. Cyanophycin synthesis was enhanced in salt-stressed cells grown in nitrate. When cells were restored to their normal growth medium the content of these substances decreased towards control levels.

Recombinant bioprocess optimization for heterologous protein production using two-stage, cyclic fed-batch culture.

Abstract: A two-stage, cyclic fed-batch bioprocess was designed, and its performance evaluated to improve rice α-amylase productivity by the yeast Yarrowia lipolytica SMY2 (MatA, ade1, ura3, xpr2), ATCC 201847, containing a replicative plasmid coding for a rice α-amlyase. Transcription of the recombinant gene is controlled by the XPR2 promoter. The first stage (or growth stage) was operated in the fed-batch mode, and the growth medium, designed to maintain a constant high cell density (i.e., 60 g/l), was fed according to a predetermined and preprogrammed optimal feed rate which, in turn, maintained the specific cell growth rate at an optimal value (i.e., 0.1 h−1). Typically, when the volume in the first stage reached a preset value, a portion of culture broth (i.e., 55%) was transferred to the second stage (or production stage). The remaining cells in the growth stage were then fed with fresh growth medium according to the bioprocess control strategy developed, while induction of α-amylase expression and its production was taking place in the second stage. The second stage was also operated in the fed-batch mode, and the production medium designed to maintain a constant high cell density and high productivity of heterologous protein was fed at a predetermined and preprogrammed rate, which maintained the specific cell growth rate at an optimal level. The volumetric α-amylase productivity achieved (1835 units l−1 h−1) from the two-stage, cyclic fed-batch culture process was twofold higher than that of the fed-batch culture process. The genetic stability of the recombinant strain and the design of optimal media for growth and production stages are also critically important to a successful implementation of the two-stage, cyclic fed-batch process for production of heterologous protein.

Influence of auxins on alkaloid accumulation by a transgenic cell line of Catharanthus roseus

Abstract: The transgenic cell line of Catharanthus roseus (L) G Don S10 was used to study the effect of the presence of the synthetic auxins naphthalene acetic acid and 2,4-dichlorophenoxyeacetic acid in the culture medium on the accumulation of terpenoid indole alkaloids Line S10 carries a recombinant, constitutively over-expressed version of the endogenous strictosidine synthase gene The experiments were carried out using a two-stage culture system, consisting of a growth phase of 7 to 10 days and a production phase of 14 or 30 days The hormonal composition was a crucial factor when formulating both the growth and the production media It was determined that the presence of naphthalene acetic acid during the production phase led to lower levels of alkaloid accumulation The presence of 2,4-dichlorophenoxyacetic acid in the growth medium reduced culture aggregation and repressed secondary metabolism Cultures grown in medium containing 2,4-dichlorophenoxyacetic acid showed reduced capacity to supply biosynthetic precursors, which resulted in low levels of accumulation of terpenoid indole alkaloids Cultures grown in 2,4-D-containing medium showed reduced capacity to supply biosynthetic precursors and higher rates of catabolic activity, which resulted in low levels of TIA accumulation The expression of the gus and strictosidine synthase transgenes, measured at the enzyme level, was similarly high under all conditions tested

Osmolarity modulates the expression of the Hha protein from Escherichia coli

Abstract: The effect of the osmolarity of the culture medium on the expression of the hha gene of Escherichia coli was investigated. When cells were grown in LB medium, expression reached a maximum in the exponential phase of growth and decreased in the stationary phase. Increasing the osmolarity of the LB medium had no significant effect on the expression of the hha gene, but depletion of NaCl led to a significant decrease in expression. Expression of the hha gene is thus sensitive to the osmolarity of the growth medium. High levels of expression of the hha gene when cells are grown at medium to high osmolarity are consistent with the finding that the Hha protein appears to play its main modulatory role when cells grow under these conditions.

Bacterial Growth Medium that Significantly Increases the Yield of Recombinant Plasmid

Abstract: Isolation of plasmid DNA from Escherichia coli is a daily activity in many molecular biology laboratories. A number of protocols and media recipes have been reported in an effort to make this process more efficient. Here we describe a growth medium that supports much higher E. coli cell densities and, concomitantly, a much higher yield of plasmid than previously reported for small-scale applications. On a unit volume basis, E. coli cultures grown in this medium, termed H15, produce up to 30-fold more recombinant plasmid than in conventional rich media, paralleling the increase in cell density. This phenomenon is independent of E. coli host strain, DNA insert size and plasmid copy number. H15 medium is also very economical; as much as 6 mg of plasmid can be harvested per dollar of medium.

The characteristics of encapsulated whole cell β-galactosidase

Abstract: We prepared encapsulated whole cell β-galactosidase using E. coli. The cell culture was divided into two steps for the cell accumulation inside the capsule and enzyme production in the cell. Growth and production media were used individually for this purpose. The dry cell weight of the free cell culture was increased 2.8 times by controlling the pH of the growth medium during cultivation. However, the weight of cells accumulated in the capsule reduced 40% with pH control. The dry cell weight increased with lactose concentration of the production medium for both cases of free and capsule cultures. The dry cell weights were 1.5 g/l for free culture and 100 g/l in the capsule when the lactose concentration of the production medium was 10 g/l. The dry cell weight increased about 60% for both cases as the lactose concentration increased from 10 to 50 g/l. The specific activity of whole cell enzyme decreased with lactose concentration from 5 to 1.4 unit/g dry cell for free culture and from 1.1 to 0.65 unit/g dry cell in the capsule. The value of Michaelis constant, Km, of whole cell enzyme increased 3 times because of the resistance of mass transfer through the capsule membrane. The constants of Michaelis-Menten equation for the whole cell enzyme in the capsule were Vm: 0.0479 mM/min and Km: 44.86 mM. These constants of the membrane-free cells were Vm: 0.0464 mM/min and Km: 15.64 mM. To increase the whole cell enzyme activity, we treated encapsulated cells with organic solvents. The activity of encapsulated whole cell enzyme was increased 3.5 times with the treatment of chloroform and ethanol. The activity of the encapsulated whole cell enzymes was reserved after repeating the process 30 times.

Hinokitiol Production in a Suspension Culture of Calocedrus formosana Florin.

Abstract: A suspension culture of Calocedrus formosana Florin was studied as a material for efficient production of hinokitiol. Murashige-Skoog's medium containing 3% sucrose and 1 mg/l 1-naphthylacetic acid was most desirable for cell growth. Cell growth, expressed as fresh cell weight, showed a 20-fold increase after 4 weeks of culture in this medium. Adding potassium acetate or chitosan to the medium increased hinokitiol production. The highest hinokitiol yield, 1700 μg/g fresh cells, was obtained when cells were cultured in the growth medium with chitosan.

Ethanol production by mutant yeast

Abstract: The invention relates to methods of producing ethanol and yeast cells that may be used according to the method. Said method comprising growing respiratory deficient yeast cells (such as K1Δpet191ab described herein) which have at least one nuclear gene, or product thereof, required for respiration which is non-functional and/or inhibited in a growth medium. The method may be used in the production of fuel ethanol or for the production of alcoholic drinks.

Medium for recovery and growth of microorganisms

Abstract: Reagents for use in the recovery and growth of microorganisms, comprise (a) growth medium generating low levels of toxic oxidising species; (b) a hydrogen donor; and (c) freeze-dried OXYRASE enzyme. The combination of use of a growth medium generating low levels of toxic oxidising species together with OXYRASE enzyme enables significantly improved recovery, i.e. better and faster recovery, of stressed microorganism cells than is possible with conventional growth media. Not only are more stressed cells recoverable, but also recovery of all cells takes place in less time than under traditional conditions.

Loss of fimbrial adhesion with the addition of vaccznum escherchia coli macrocarpon to the growth medium of p-fimbriated

Abstract: Purpose: Vuccinium macrocarpon-the American cranberry-irreversibly inhibits the expression of P-fimbriae of E. coli. Further effects on the function and expression of P-fimbriae were studied by growing P-fimbriated E. coli in solid media laced with cranberry juice. Methods: Cranberry concentrate at pH 7.0 was added to CFA medium to a final concentration of 25%. E. coli strains JR1 and DS17 were plated on this medium with a plain CFA control and incubated at 37C. Cultures were tested for ability to agglutinate P-receptor specific beads. Bacteria were washed in PBS and agglutination retested. Cultures were also replated on plain CFA agar and rechecked for their ability to agglutinate. Transmission electron micrographs were performed on positive control and test bacteria. Results: For E. coli strain JR1, P-fimbrial agglutination was inhibited after the third plating. DS17 was fully inhibited after the second plating. Washing in PBS did not affect agglutination, but replating on CFA agar allowed agglutination to recur. Electron micrographic study of control populations confirmed fimbriae. Fully inhibited bacteria had a 100% reduction in expression of fimbriae. Additionally, inhibited bacteria showed cellular elongation. Conclusions: Cranberry juice irreversibly inhibits P-fimbriae. Electron micrographic evidence suggests that cranberry juice acts on the cell wall preventing proper attachment of the fimbrial subunits or as a genetic control preventing the expression of normal fimbrial subunits or both.

Formation and Regeneration of Protoplasts from Paecilomyces Cicadae

Abstract: The effects of such factors as enzyme systems, media for mycelia growth, osmotic stabilizers, different temperatures and age of mycelium on the formation of the mycelia protoplasts were studied.High yield of protoplasts were obtained from 16 hours mycelia of Paecilomyces cicadae by using 1.5% lyvallzyme as enzyme system, starch agar medium as mycelia; growth medium and 0.6% KCl as osmotic stabilizer at the temperature of 28℃. When 0.6M NaCl was used as stabilizer, about 7.97% of protoplasts regenerated and formed colonies on soybean medium (SOS). The morphological processes of protoplasts and regeneration were illustrated by microphotograph.

Increasing Monoclonal Antibody Productivity by Semicontinuous Substitution of Production Medium for Growth Medium

Abstract: 1. A hybridoma perfusion culture can be maintained at steady state (viability >90%, cell density up to 7x106cells/ml) for arbitrary time period, thanks to the enrichment of the medium with apoptosis-suppressing amino acids, and to the application of a high cell bleed rate. 2. The perfusion cell growth modul with a working volume of 1.0 1 can serve as a source of ∼2.0x109cells available daily for initiation of short-term batch cultures. 3. Due to high initial cell density, and substantial exhaustion of nutrients in the growth medium, the batch cultures behave as if initiated at their stationary phase. 4. If the medium is sufficiently enriched with the building stones of the Mab molecule, i.e. the essential amino acids, and with the main energy source, glutamine, the synthesis of Mabs proceeds at high specific rate. The exhaustion of glutamine, but not of glucose, acts as the stop-signal for Mab synthesis. 5. The Mab concentration in the medium harvested from the batch cultures is several-fold higher than in the medium harvested directly from the perfusion cell growth modul.

Method of incorporating amino acid analogs using a hypertonic growth medium

Abstract: Incorporation of certain amino acid analogs into polypeptides produced by cells which do not ordinarily provide polypeptides containing such amino acid analogs is accomplished by subjecting the cells to growth media containing such amino acid analogs. The degree of incorporation can be regulated by adjusting the concentration of amino acid analogs in the media and/or by adjusting osmolality of the media. Such incorporation allows the chemical and physical characteristics of polypeptides to be altered and studied.