Preparation and characterization of calibration beads for sorting cells expressing a beta-lactamase gene reporter.

BACKGROUND: Modern drug discovery has been based on high-throughput screening using whole-cell assays. A prominent role has been assigned to the reporter gene technology based on a beta-lactamase and the fluorogenic substrate CCF2. Successful application of this technology requires fluorescence-activated cell sorting. We describe the preparation and characterization of calibration beads for sorting cells expressing the beta-lactamase gene using the CCF2 substrate. METHODS: To model Forster resonance energy transfer (FRET) between the coumarin donor and the fluorescein acceptor of the CCF2 reporting dye, we used activated polystyrene beads with primary amino groups. Donor and acceptor fluorophores were attached to the beads at different ratios via succinimidyl esters. The beads were characterized with a fluorescence plate reader and a flow cytometer. RESULTS: We prepared polystyrene beads with five different ratios of donor and acceptor fluorophores and beads that carried a donor or a receptor fluorophore alone. Fluorescence measurements demonstrated that the prepared beads well represent the FRET of CCF2 substrate. CONCLUSION: We have demonstrated that the prepared beads can be successfully used for the setup of fluorescence-activated cell sorting to sort cells with CCF2 reporter substrate and the beta-lactamase reporter gene.     

Über die Wirkung der Quarzlampenbestrahlung der Versuchstiere auf die Autohämolyse in Vitro

Ohne Zusammenfassung     

Effects of plant density and nutrient levels on fruit polymorphism in Atriplex sagittata

Atriplex sagittata is a heterocarpic species producing three types of fruits which differ in morphology and ecological properties. This study focused on variation in biomass allocation into particular fruit types under different density and fertilization levels. The reduction in total weight under stressful conditions was accompanied by a reduction of reproductive structures in terms of both total fruit production and mean fruit weight. Allocation of biomass to particular fruit types under different density-fertilization treatments showed considerable variation. The non-dormant, bracteolate fruit type (termed C) contributed 80–90% to the total fecundity and its production was not affected by density-fertilization treatments. The production of this fruit type is higher in the upper part of the maternal plant stem. Production of the very dormant ebracteate fruit type (termed A) increased under favourable conditions and was greater on the lower part of the stem, whereas more of the so-called type B fruit (covered by bracteoles and exhibiting dispersal and dormancy characteristics intermediate between types A and C) was produced under suboptimal conditions and was concentrated in the middle part of the plant body. This pattern contradicts to some extent the theory that deeply dormant and less dispersible fruits will be produced under stressful conditions and may be considered a new model for the behaviour of plants with more than two heterocarpic fruits. The type B fruit, intermediate in both morphology and ecological behaviour, provides a continuum between extreme strategies. Received: 8 February 1998 / Accepted: 19 November 1998     

How does seed heteromorphism influence the life history stages of Atriplex sagittata (Chenopodiaceae)?

Atriplex sagittata is an annual, heterocarpic species producing three types of fruits, (i) dormant ebracteate (further termed type A), (ii) dormant bracteate (type B) and (iii) non-dormant bracteate (type C). In this study, we investigated populations grown from particular fruit types under different density regimes, and assessed their fate in particular life stages throughout a complete growing season. Differences in germination date among populations grown from seeds of the three fruit types were found, followed by differences in population density and plant size. Low population density and total biomass was recorded in populations of types A and B fruit (which are smaller and exhibit less germination), but the low density was not associated with faster growth of individual plants. Type C fruit, with bigger seeds and higher germinability, generated populations with higher densities and higher total biomass than populations generated from type A and B. There was a significant effect of plant density on dry weight of individual plants, regardless of the fruit type from which they were grown. Under both density regimes, plants grown from types A and B fruit were lighter than those grown from type C fruit. Plants grown from different fruit types produced dispersal units (fruits+bracteoles) of different mean weight which increased from A to B to C. However, the total fruit weight was not significantly different when plant size was used as a covariate. Maternal effect on seed germinability, exhibited by plants grown from the three fruit types, was not significant. However, a remarkable trend was found. Lower germination was recorded in type A and B fruits produced by plants which grew under higher densities. It is concluded that plants derived from different fruit types differ in growth characteristics and population parameters which can have pronounced effect on the life cycle and population regeneration in successive years.     

Towards a better understanding of the Chenopodium album aggregate (Amaranthaceae) in the Middle East: A karyological,cytometric and morphometric investigation

The study of variation in nuclear genome size, especially when combined with common garden experiments, significantly contributes to disentangling interspecies relationships within taxonomically complicated plant groups. The Chenopodium album aggregate is among the morphologically most variable groups and consists of many weakly differentiated cosmopolitan entities. We analysed nuclear genome size variation in diploid and polyploid species of the aggregate from Iran using flow cytometry of 282 accessions from 88 populations of 7 species. To this end, we also determined chromosome numbers and performed a morphometric study to reveal the extent of intraspecific morphological variation. We found that Iranian species are exclusively diploid (C. vulvaria), tetraploid (C. novopokrovskyanum, C. strictum, C. sosnowskyi and C. chaldoranicum) or hexaploid (C. album subsp. album, C. album subsp. iranicum and C. opulifolium). Six homogeneous relative genome size groups were distinguished among the species studied. Our morphometric study surprisingly revealed that under similar ecological conditions Chenopodium species are morphologically stable and well distinguishable, exhibited very little morphological variation. Hence, immense variation in leaf shapes, branching and inflorescence organization seen in the field has not been repeated under greenhouse conditions. The only exception was C. album s. str. which exhibited numerous morphotypes, covering the variation of remaining species.     

Preparation and Enantioselectivity Binding Studies of a New Chiral Cobalt(II)porphyrin‐Tröger's Base Conjugate

A new bis[cobalt(II)porphyrin]‐Tröger's base conjugate was studied as a potential receptor for methyl esters of several amino acids. The conjugate was prepared as racemate, and then resolved via preparative high‐performance liquid chromatography (HPLC) on a chiral column. The high affinity to lysine, histidine, and proline methyl esters was found by complexation studies followed by UV‐Vis spectroscopy. The studies of pure enantiomers, followed by UV‐Vis and electronic circular dichroism spectroscopy, revealed the highest enantioselectivity for lysine methyl ester. Chirality 26:361–367, 2014. © 2014 Wiley Periodicals, Inc.     

Epidemiology of Helicobacter pylori infection in the Czech Republic

BACKGROUND: Prevalence of Helicobacter pylori infection has been estimated to range from 60 to 95% in the former communist countries of Central and Eastern Europe. The aim of this study was to evaluate H. pylori infection prevalence in a representative sample of the Czech population. The second objective was to describe difference of H. pylori prevalence between different social groups of children and adults. MATERIALS AND METHODS: A total of 2509 persons aged 5-100 years, randomly selected out of 30,012 persons of the general population, took part in the study. H. pylori infection was investigated by means of 13C-urea breath test. Breath samples were analyzed by isotope ratio mass spectrometry. Social and demographic characteristics were based on data from self-completed questionnaires. RESULTS: Using the total Czech population as a standard, we estimated the age-standardized prevalence of H. pylori in males aged 5+ years at 41.9% (95% CI 39.0%, 44.8%) and in females aged 5+ years at 41.4% (95% CI 38.6%, 44.3%) in 2001. Prevalence of H. pylori increased with age but was not related to gender. Children of mothers with basic or lower education, living in crowded accommodations, without access to running warm water, and residing in smaller towns appear to be at the highest risk. Low education and heavy smoking are most strongly associated with prevalence of H. pylori positivity in adults and adolescents. CONCLUSIONS: This is a unique study based on a representative sample of the general population in a Central European country. The overall prevalence of H. pylori is lower than previously assumed and could partly reflect a substantial recent decrease in H. pylori prevalence in the Czech Republic. Consistent with earlier studies, H. pylori infection is strongly influenced by socioeconomic conditions and childhood poverty.