Restriction enzyme cleavage of fluorescently labeled DNA fragments--analysis of the method and its usage in examination of digestion completeness

Restriction enzymes have proven to be among the most valuable tools in molecular biology. In this work, we demonstrate that the cleavage of fluorescently labeled, PCR-amplified DNA can be used as a simple and highly sensitive technique for detection of sequences present in a percentage as low as 0.6% in a DNA pool. Due to the fact that fluorescent labeling of DNA fragments enables such sensitive detection and quantification of restriction enzyme cleavage, the method was further exploited in monitoring of the enzymatic digestion completeness and in determination of factors that influence restriction enzyme effectiveness. We analyzed the activity of six restriction endonucleases; the percentage of uncleaved DNA fragments predominantly ranged between 2.0 and 2.5 and the highest value was 8.00%. We conclude that, since the enzymatic digestion completeness may not always be assured, each assay based on restriction enzyme cleavage that is intended to be used in investigations of heterogeneity in a DNA pool should be constructed so that the presence of cleaved sequences is the indication of pool nonuniformity. When the presence of uncleaved sequences indicates pool heterogeneity, the results could be misleading due to possible incompleteness of enzymatic cleavage.     

Beneficial effect of agmatine in the acute phase of experimental autoimmune encephalomyelitis in iNOS-/- knockout mice

The aim of the study was to investigate the hypothesis that agmatine (AGM) provides protection against oxidative stress in experimental autoimmune encephalomyelitis (EAE).     

Population dynamics of mangrove Avicennia bicolor on the Pacific coast of Costa Rica

Deterministic and stochastic class structured population models were used to simulate the life cycle of Avicennia bicolor of the Pacific coast of Costa Rica. The models were based on an extensive data set collected during a 6 year period in a 0.52 ha plot of monospecific A. bicolor. This data set included density, growth, mortality and transition rates of seedlings, saplings and trees of eight different diameter classes, as well as propagule production for the reproductive tree classes. Model simulations carried out over a 100 year period indicated a stable size class structure of the forest. Sensitivity analysis showed a significantly greater sensitivity of the model population to simulated changes in the mortality of seedlings, in comparison with the mortality of saplings and trees. An increase of 1% in the mortality of seedlings, for example, was sufficient to cause significant changes in the density of individual size classes. In contrast, neither a 10% increase in the mortality of saplings and trees nor a 20% decrease in the propagule production of fecund trees significantly affected the overall forest structure.