Growth medium

About: Growth medium is a research topic. Over the lifetime, 1889 publications have been published within this topic receiving 59171 citations. The topic is also known as: culture medium & culture media.

Factors affecting the electroporation of Bacillus subtilis

Abstract: Bacillus subtilis 168 trp- was found to be transformable with the tetracycline resistance plasmid pAB124 by electroporation of whole cells, inconsistently and at very low frequencies. Supplementation of the growth medium with glycine, or particularly DL-threonine, produced cells that could be electrotransformed much more efficiently at frequencies up to 2.5 x 10(3) transformants per microgram plasmid DNA. Transformation was optimal with cells grown in medium containing a racemic mixture of the D- and L-isomers of threonine, and no transformants were obtained when pure forms of the D- and L-threonine isomers were used. The cell walls of B. subtilis grown in the presence or absence of D-, L- and DL-threonine had a similar amino acid composition which did not include threonine. A more complex biochemical explanation of the enhancement of electroporation by growth in DL-threonine is likely, and this is discussed. Lysozyme treatments to weaken the cell wall and possibly mimic the effect of DL-threonine did not yield any transformants. The effects of buffer composition and culture incubation time were also determined and the electroporation protocol optimized accordingly. The response of a range of other B. subtilis strains to electroporation by the method produced was found to be variable. In all cases, transformation was verified by recovery of the plasmid DNA from putative transformants.

β-glucosidase production by Trichoderma reesei

Abstract: The hydrolysis of cellulose to the water-soluble products cellobiose and glucose is achieved via synergistic action of cellulolytic proteins. The three types of enzymes involved in this process are endoglucanases, cellobiohy-drolases, and β-glucosidases. One of the best fungal cellulase producers is Trichoderma reesei RUT C30. However, the amount of β-glucosidases secreted by this fungus is insufficient for effective cellulose conversion. We investigated the production of cellulases and β-glucosidases in shake-flask cultures by applying three pH-controlling strategies: (1)the pH of the production medium was adjusted to 5.8 after the addition of seed culture with no additional pH adjustment performed, (2) the pH was adjusted to 6.0 daily, and (3) the pH was maintained at 6.0 by the addition of Tris-maleate buffer to the growth medium. Different carbon sources-Solka Floe 200, glucose, lactose, and sorbitol—were added to standard Mandels nutrients. The lowest β-glucosidase activities were obtained when no pH adjustment was done regardless of the carbon source employed. Somewhat higher levels of β-glucosidase were measured in the culture filtrates when daily pH adjustment was carried out. The effect of buffering the culture medium on β-glucosidase liberation was most prominent when a carbon source inducing the production of other cellulases was applied.

Predisposition of Cloned Fetal Hamster Lung Epithelial Cells to Transformation by a Precarcinogen, Benzo(a)pyrene, Using Growth Hormone Supplementation and Collagen Gel Substratum

Abstract: A cloned fetal Syrian hamster lung epithelial cell line (M3E3/C3) was used to compare the influence of two different culture conditions on the degree of cellular differentiation and susceptibility of the cells to undergo malignant transformation by a precarcinogen, benzo(a)pyrene. Conventional conditions consisted of growth medium containing Roswell Park Memorial Institute Medium 1640, pyruvate, and fetal bovine serum and a substratum of plastic. Complex conditions comprised the growth medium supplemented with insulin, hydrocortisone, estradiol, epidermal growth factor, transferrin, and cholera toxin and a substratum of collagen gel. Under the complex culture conditions, there was extensive development of endoplasmic reticulum and Golgi vesicles, whereas under conventional conditions these organelles were only minimally developed. This was correlated with 1.5-1.8 times enhancement of ethoxycoumarin deethylase and reduced nicotinamide adenine dinucleotide phosphate-dependent cytochrome c reductase activities. Decomposition of added benz(a)anthracene into water-soluble compounds increased with the period of incubation and reached about 40% of initial benz(a)anthracene (50 micrograms/10 ml/flask) at 48 h under the complex conditions, whereas under the conventional conditions only less than 4% decomposition occurred. Benzo(a)pyrene in the dose range 2-8 micrograms/ml was strongly cytotoxic and caused significant anchorage independent transformation only under complex culture conditions. Transformed cells produced tumors in two of four hamsters during 8 months following s.c. injection within 48 h of birth. These results suggest that the complex culture conditions predisposed the cloned fetal epithelial cells to malignant transformation by benzo(a)pyrene through stimulation of cellular differentiation and development of enzyme systems capable of activating it metabolically.