A high efficiency method for site-directed mutagenesis with any plasmid

A procedure is described which allows for the site-directed mutagenesis of DNA segments in any double-stranded plasmid with high efficiency. There are no limitations as to the position of the mutation. The protocol involves only simple enzymatic manipulations and no difficult to control operations, such as partial digestions, are required. The method was developed and used to mutagenize two different genes (encoding human interferon-beta and interleukin-2) cloned in a eukaryotic expression vector. For ten mutageneses with different oligodeoxyribonucleotides the average yield of mutants was 60%.     

Differential behavior of two dexamethasone induced mRNA activities in HTC cells in response to cordycepin and to withdrawal of hormone.

In HTC cells dexamethasone induces the accumulation of tyrosine aminotransferase mRNA activity in addition to a mRNA activity coding for a protein of a yet unknown function (DIP). The kinetics of decay of these two mRNA activities were followed in induced cells under the influence of cordycepin and in cells withdrawn from the hormone. For tyrosine amino-transferase mRNA activity a relative half life of approximately 1 hour was estimated in both cases. However, DIP mRNA activity decayed to 50 % of the fully induced level within 3 to 4 hours following withdrawal of the hormone, whereas after application of cordycepin to induced cells only a small and delayed decay could be observed. Application of cordycepin two hours after withdrawal of hormone caused a stabilization of DIP mRNA activity.     

Metabolic heterogeneity of muscle fibers classified by myosin ATPase

Summary Muscle fibers re commonly classified histochemically into three types by the staining intensity for myosin ATPase combined with those for metabolic enzymes. Preincubation at pH 4.6 gives rise to three staining intensities of myosin ATPase which are also used for fiber typing. The two classification systems were compared by computer analysis of the individual staining profiles of over 2,500 fibers, and found not to be equivalent. The analysis showed metabolic heterogeneity among the fiber groups distinguished according to their differences in myosin ATPase.     

Effects of domestic wastewater on serum enzyme activities of brown trout (Salmo trutta)

1. Chronic injuries of kidney and liver tissue of brown trout caused by domestic wastewater were not accompanied by increased activities of serum enzymes (GOT, GPT). 2. Increased enzyme activities were only observed in fish with acute Saproloegnia infection. 3. It is important to distinguish between acute and chronic responses of serum enzymes in diseased fish.     

Genetic parameters of a random regression model for daily feed intake of performance tested French Landrace and Large White growing pigs

Daily feed intake data of 1 279 French Landrace (FL, 1 039 boars and 240 castrates) and 2 417 Large White (LW, 2 032 boars and 385 castrates) growing pigs were recorded with electronic feed dispensers in three French central testing stations from 1992–1994. Male (35 to 95 kg live body weight) or castrated (100 kg live body weight) group housed, ad libitum fed pigs were performance tested. A quadratic polynomial in days on test with fixed regressions for sex and batch, random regressions for additive genetic, pen, litter and individual permanent environmental effects was used, with two different models for the residual variance: constant in model 1 and modelled with a quadratic polynomial depending on the day on test d_m as follows in model 2: . Variance components were estimated from weekly means of daily feed intake by means of a Bayesian analysis using Gibbs sampling. Posterior means of (co)variances were calculated using 800 000 samples from four chains (200 000 each). Heritability estimates of regression coefficients were 0.30 (FL model 1), 0.21 (FL model 2), 0.14 (LW1) and 0.14 (LW2) for the intercept, 0.04 (FL1), 0.04 (FL2), 0.11 (LW1) and 0.06 (LW2) for the linear, 0.03 (FL1), 0.04 (FL2) 0.11 (LW1) and 0.06 (LW2) for the quadratic term. Heritability estimates for weekly means of daily feed intake were the lowest in week 4 (FL1: 0.11, FL2: 0.11) and week 1 (LW1: 0.09, LW2: 0.10), and the highest in week 11 (FL1: 0.25, FL2: 0.24) and week 8 (LW1: 0.19, LW2: 0.18), respectively. Genetic eigenfunctions revealed that altering the shape of the feed intake curve by selection is difficult.     

Frequency of pathogenic enterobacteria in infantile diarrheic processes in the city of Recife, Pernambuco, Brazil

326 samples of diarrheal feces obtained from children whose ages ranged from zero to 5 years, admitted in two rehydration hospitals in the city of Recife, Pernambuco, were analyzed. Feces were placed in Cary-Blair medium (4 degrees C) for shipment to the laboratory. There was no difference in the rate of bacteria isolation if the samples were analyzed within the period from 3 to 7 days of collection. 19.02% of the analyzed samples were positives for at least one of the searched bacteria, 26 Salmonella belonging to 3 species, 21 classic enteropathogenic E. coli, 1 invasive E. coli, 10 Shigella belonging to 3 serotypes and 1 Yersinia enterocolitica were found.     

The adaptation of digestive enzymes to temperature, season and diet in roach Rutilus rutilus L. and rudd Scardinius erythrophthalmus L. Cellulase

The cellulolytic activity of the contents of the intestinal tract of populations of roach and rudd in four Tyrolean lakes was studied. Much of the activity was taken up with the diet. In fish feeding on plants and detritus a lower cellulolytic activity was found than in fish feeding on zooplankton or on arthropods. A seasonal pattern of cellulolytic activity was demonstrated with a peak in the autumn. It is assumed that the cellulolytic activity found in the intestinal contents of cyprinids is insufficient for the digestion of fibre, but that is suffices—particularly in omnivorous species—to support the breakdown of cell walls.