Hybridase cleavage of RNA. I. Oligonucleotide probes for regiospecific RNA cleavage]

New oligonucleotide probes for regiospecific cleavage of RNA molecules by hybridase (RNase H) are suggested. RNase H from E. coli is shown to site-specifically split eight phosphodiester bonds in RNA in the heteroduplex, formed by 5S rRNA and d(ACCACCGCGCT). The partial substitution of deoxycytidines in position 5, 6, 8, 10 of the probe by 2'-O-methylcytidines leads to unique (regiospecific) RNA cleavage between U25 and C26.     

Taxonomic spectrum of pathogens of mixed infections in oncology patients and principles of their antibacterial therapy

Mixed infections were observed in oncological patients irrespective of the tumor or infectious process localization. 25-30 per cent of the pathological materials from such patients were contaminated with bacterial associations. The number of the associates ranged from 2 to 5 depending on the pathological material tested. Therapy of infectious complications due to pathogen associations should stem from analysis of many factors, i.e. properties of the isolated or likely pathogens, individual characteristics of the patients, compatibility of the drugs used and their pharmacokinetic features.     

A karyological study of Asphodelus L. (Asphodelaceae) from the Western Mediterranean

The following aspects of Asphodelus karyology are analysed: base number, polyploidy, chromosome size, chromosome morphology, satellited chromosomes, structural heteromorphism, karyotype asymmetry and karyotype evolution. The base number 0 ×= 14 is common to all species except for A. refractus , which has the derived ×= 13. Three ploidy levels occur, often in the same species; diploid, tetraploid and hexaploid, with 2n = 28, 56 and 84. Chromosomes are generally small to medium-small, with the occasional presence of medium-large chromosomes. The most frequent chromosome types are metacentric of type m and submetacentric. Metacentric chromosomes of type M occur only in sections. Verineopsis, Verinea and Plagiasphodelus ; subtelocentric chromosomes occur only in sections Asphodelus and Plagiasphodelus. There is a wide variability in relation to the number of satellited chromosomes, relative to ploidy level. There are usually two to four in diploids, four to eight in tetraploids and usually six, exceptionally up to 12, in the hexaploid. Satellites are present on the shortest arm, exceptionally on the longest arm. There is a high degree of structural heteromorphism in practically all the species which affects satellited and non satellited chromosomes. Karyotype asymmetry is generally of type 2B. Inter-and intra-chromosomal differences are estimated by the A1 and A2 indexes. Both indices vary in the karyotype evolution of the genus, with a decrease of A1 and an increase of A2. The role of polyploidy, hybridization, asymmetry and decrease of chromosome size in the evolution of Asphodelus is discussed.     

Somaclonal variation as a tool to develop pest resistant plants of Torenia fournieri ‘Compacta Blue’

Callus cultures of Torenia fournieri Compacta Blue were initiated on a modified Murashige and Skoog salt medium (MS) with 2.26 M 2,4-dichlorophenoxyacetic acid. Shoots were regenerated from these cultures using MS medium amended with 2.46 M indolebutyric acid and 8.88 M benzyladenine. These shoot cultures were subjected to two-spotted spidermite (Tetranychus urticae Koch.) and the greenhouse whitefly [Trialeurodes vaporariorum (Westwood)]. Pests were allowed to feed until such time that their populations started to decrease due to lack of food. The remaining live tissue of the Torenia was placed on MS medium amended with 2.28 M zeatin to induce new adventitious shoots and plantlets. Newly regenerated plantlets were acclimated to greenhouse conditions and evaluated for resistance to the pest to which they were subjected in vitro. Highly significant differences in pest numbers were found in somaclones for both the two-spotted spidermite and greenhouse whitefly when compared to control plants. A wide range of variability was observed among the somaclonal population. There were significantly fewer mite eggs laid on plants regenerated from in vitro cultures screened with two-spotted spidermites than on seed sown controls. Regenerants from cultures screened with whiteflies in vitro had fewer eggs, immatures and live adults than controls.Abbreviations BA benzyladenine - IBA indolebutyric acid - 2,4-d 2,4-dichlorophenoxyacetic acid - MS Murashige and Skoog salt medium Storrs Agricultural Research Station Scientific Publication 1641.     

Gas exchange in leaves of the root hemiparasitemelampyrum arvense L. before and after attachment to the host plant

Gas exchange characteristics of a hemiparasiteMelampyrum arvense L. before and after attachment to the hostCapsella bursa pastoris (L.) Med. were compared. The net photosynthetic rates (P_N) on a leaf area basis were extremely low and in comparison to the value obtained for the host were about 15 % and 23 % for the unattached and attached hemiparasite, respectively. Also the concentration of photosynthetic pigments was low (as compared with the host the content of chlorophylls was about 33 % and 49 % and of carotenoids about 38 % and 36 % in the unattached and attached hemiparasite, respectively). On the other hand the rates of respiration were high (about 1.8 and 2.6 times higher in the unattached and attached hemiparasite, respectively, than in the host). In darkness stomatal conductance (g_S) of the host and the unattached hemiparasite was rapidly reduced to 10 % of the value obtained in light, g_S of the attached hemiparasite was decreased only by about 30%. A total reduction of g_S occurred at relative water content (RWC) of 85 %, 75% and 45 % for the unattached hemiparasite, the host, and the attached hemiparasite, respectively. The transpiration (E) rate in the preparasitic stage was very low, being 2.6 and 4.5 times smaller than in the host and the attached hemiparasite, respectively. In the attached hemiparasite WUE was 7.5 and 3 times poorer than in the host and in the preparasitic stage, respectively.     

Planar asymmetric lipid bilayers of glycosphingolipid or lipopolysaccharide on one side and phospholipids on the other: membrane potential, porin function, and complement activation.

We have determined some physicochemical properties of the monosaccharide-type fraction (GSL-1) of glycosphingolipids, the major glycolipid components of the outer leaflet of the Gram-negative species Sphingomonas paucimobilis. These properties included the state of order of the hydrocarbon moiety, the effective molecular area, surface charge density, and intrinsic transmembrane potential profile of reconstituted planar asymmetric GSL-1/phospholipid bilayer membranes. We have, furthermore, investigated the insertion into and the function of porin channels in the reconstituted bilayers and the complement-activating capability of GSL-1 surfaces. All results were compared with respective data for deep rough mutant lipopolysaccharide of Salmonella minnesota R595. We found a remarkable agreement in most functional properties of the two glycolipids.     

Electrophysical monitoring of culture process of recombinant Escherichia coli strains

A novel method for monitoring the cell culture process has been developed. The method is based on the measurements of electro-optical characteristics of cell suspension, calculation of cell structure parameters, and the relationship between accumulation of proteins and change of these parameters' employment. Application of the method for the monitoring of a culture process of a recombinant strain is considered. The process of growth of recombinant strains cannot be sufficiently predicted and the direct measurement of cell culture parameters is unlikely to be the most efficient way of solving the problem.Escherichia coli plasmid-free and recombinant strains synthesizing the fusion protein consisting of tumor necrosis factor-alpha (TNF) and thymosin-alpha(1) (T) were studied. It was found that cytoplasmic electroconductivity of the strains investigated increased during the culture process. The accumulation of insoluble recombinant pThy-315-encoded hybrid protein TNF(SINGLEBOND)T in cells resulted in a decrease of the membrane dielectric permeability. To determine variations of membrane dielectric permeability the amount of insoluble recombinant protein TNF(SINGLEBOND)T in the bacterial cells should be calculated.     

Approximate methods for estimating the pattern of nucleotide substitution and the variation of substitution rates among sites

We propose two approximate methods (one based on parsimony and one on pairwise sequence comparison) for estimating the pattern of nucleotide substitution and a parsimony-based method for estimating the gamma parameter for variable substitution rates among sites. The matrix of substitution rates that represents the substitution pattern can be recovered through its relationship with the observable matrix of site pattern frequences in pairwise sequence comparisons. In the parsimony approach, the ancestral sequences reconstructed by the parsimony algorithm were used, and the two sequences compared are those at the ends of a branch in the phylogenetic tree. The method for estimating the gamma parameter was based on a reinterpretation of the numbers of changes at sites inferred by parsimony. Three data sets were analyzed to examine the utility of the approximate methods compared with the more reliable likelihood methods. The new methods for estimating the substitution pattern were found to produce estimates quite similar to those obtained from the likelihood analyses. The new method for estimating the gamma parameter was effective in reducing the bias in conventional parsimony estimates, although it also overestimated the parameter. The approximate methods are computationally very fast and appear useful for analyzing large data sets, for which use of the likelihood method requires excessive computation.