In vitro culture selection increases glyphosate tolerance in barley

In vitro culture of barley calluses has been used to produce plants with increased glyphosate tolerance. Calluses from immature embryos of barleyHordeum vulgare L. (Jeff) were cultured on Murashige and Skoog medium with 10^-6, 10^-5, 10^-4, 5×10^-4, 10^-3, or 10^-2M glyphosate for one, four or thirty months. Plants were regenerated from calluses maintained in glyphosate medium at 10^-6, 10^-5 or 10^-4M for four months, at 10^-5 or 5×10^-4M for one month and at 10^-5M for thirty months. The progeny of each regenerated plant was analyzed for response to glyphosate. Some progenies showed increased tolerance to glyphosate.     

Some events of mitosis and cytokinesis in the generative cell of Ornithogalum virens L.

The organization of the microtubule (Mt) cytoskeleton during mitosis and cytokinesis of the generative cell (GC) in Ornithogalum virens L. (bicellular pollen type, chromosome number, n = 3) from prophase to telophase/sperm formation was investigated by localization of -tubulin immunofluorescence using a conventional fluorescence microscope and a confocal laser scanning microscope. Chromosomes were visualized with DNA-binding fluorochrome dyes (ethidium bromide and 46-diamino-2-phenyl-indole). The GC of O. virens is characterized by G2/M transition within the pollen grain and not in the pollen tube as occurs in the majority of species with bicellular pollen. It was found that prophase in the GC starts before anthesis and prometaphase takes place after 10 min of pollen germination. The prophase Mts are organized into three prominent bundles, located near the generative nucleus. The number of these Mt bundles is the same as the number of GC chromosomes, a relation which has not previously been considered in other species. The most evident feature in the prophase/ prometaphase transition of O. virens GC is a direct rapid rearrangement of Mt bundles into a network which appears to interact with kinetochores and form a typical prometaphase Mt organization. The metaphase chromosomes are arranged into a conventional equatorial plate, and not in tandem as is thought to be characteristic of GC metaphase. The metaphase spindle consists of kinetochore fibres and a few interzonal fibres which form dispersed poles. Anaphase is characterized by a significant elongation of the mitotic spindle concomitant with the extension of the distance between the opposite poles. At anaphase the diffuse poles converge. Cytokinesis is realized by cell plate formation in the equatorial plane of the GC. The phragmoplast Mts between two future sperm nuclei appear after Mts of the mitotic spindle have disappeared.Abbreviations DAPI 46-diamino-2-phenyl-indole - GC generative cell - GN generative nucleus - Mt microtubule This research was made possible in part due to TEMPUS Programme and Global Network for Cell and Molecular Biology UNESCO grants to Magorzata Bana. The experimental part of the work was done in Siena University. M. Banas is very grateful to Prof. Mauro Cresti and his group for scientific interest, offering the excellent laboratory facilities, and kind reception.     

Sex influence on recombination frequency in Secale cereale L.

The variation in recombination frequency (rf) is important to plant breeders since their major objective is to obtain favorable recombinants of linked genes. One source of variation in rf is sex. Sex differences for recombination frequencies were studied in four of the seven chromosomes of Secale cereale L. cv Ailés using isozyme and storage protein loci and were determined on the basis of reciprocal crosses between heterozygous plants of cv. Ailés and homozygous plants of the inbred line Riodeva. The differences were found to be strongly segmentspecific. In some cases the level of crossing-over in male and female meiosis was about the same (between Pgm1 and Ndh1 loci on chromosome arm 4RS). However, for most of the chromosome segments in 1R, 3RL and 6RL the male rf was significantly higher than the female rf. Different hypotheses about the mechanisms of plant sex differences for recombination are discussed.     

An inquiry into the selective protection of glycine during the radiolysis of glycine-alanine mixtures in aqueous solutions and its implications to the preservation of optically active amino acids in the early earth

Akaboshi et al. (1990) has found an unexpected protection of the achiral amino acid, glycine, towards ionizing radiation at the expense of the selective destruction of the chiral amino acids, alanine and aspartic acid. The present work examines the mechanism of this protection for the case of alanine. We have developed a computer model for the radiolysis of glycine, alanine and glycine-alanine mixtures in aqueous solution. It is established that this protection is due in part to the reaction of the α-radical of glycine with alanine to regenerate a more stable α-radical, according to the following reaction, $$ \cdot CH(NH_3^ + )CO_2^ - + CH_3 CH(NH_3^ + )CO_2^ - \to CH_2 (NH_3^ + )CO_2^ - + CH_3 \dot C(NH_3^ + )CO_2^ -$$ The rate constant of this reaction was estimated to be ≤10⁴M^-1s^-1. The implications for this selective protection of glycine are considered for a hypothetical case in which there would be an enrichment of about 10% ofL-alanine in the primitive ocean and taking the glycine/alanine ratios obtained in CH₄-and CO₂- dominated atmospheres using electric discharge experiments. It is predicted that alanine would be rapidly destroyed and radioracemized in spite of the fact that the concentration of alanine is equal or significantly lower than that of glycine. Assuming that chiral amino acids were a prerequisite for the origin of life, it can be deduced that life could have appeared in a relatively short period of time unless there was a constant supply of optical amino acids from extraterrestrial sources.     

A rapid urease test for presumptive identification of Cryptococcus neoformans

A rapid method to evidence urease activity is described. Urea hydrolysis and consequent production ammonia are detected by a chemical reaction producing a blue phenol compound (indophenol blue). Three hundred and three yeast were tested. Out of 107 urease-positive organisms detected by Christensen's Urea Agar Test (CUAT) 102 were positive by our method. No false negatives were observed by this method when testing 87 Cryptococcus strains. Ths practical screening test for presumptive identification of Cryptococcus neoformans is simple, unaffected by pH changes and requires 15 minutes to be performed.     

Candida albicans stress mannoproteins expression in superficial and systemic candidiasis

The presence of heat shock mannoproteins (HSMPs) reactive with sIgA was demonstrated in several C. albicans strains. The subculture of the C. albicans isolated from mucosal surfaces on Sabouraud's dextrose agar at 25 °C switched off the HSMP expression. A re-expression of the HSMPs was obtained in the same medium by shifting the temperature of incubation to 37 °C. However, expression of HSMPs in two strains isolated from deep infections was maintained during several subcultures on Sabouraud's dextrose agar at 25 °C. A glycoprotein of 200 kDa seemed to be the main HSMP reacting with vaginal sIgA. The data presented in this study suggest that factors other than temperature can influence the expression of C. albicans HSMPs and therefore these antigens should be referred as stress mannoproteins.Abbreviations HSMPs heat shock mannoproteins - MAb monoclonal antibody - sIgA secretory IgA     

Characterization of homologous DNA recombination activity in normal and immortal mammalian cells.

Homologous DNA recombination levels were measured in normal and spontaneously immortalized murine and human fibroblasts, and in a number of primate and murine established fibroblast cell lines. Immortal cell lines and tumor-derived clones homologously recombined extrachromosomal plasmid substrates at frequencies approximately 100-fold higher than did normal cells. To further explore the mechanism responsible for this phenotype, homologous recombination frequency was measured using nuclear extracts derived from normal and immortalized murine and human fibroblasts. Extracts prepared from immortal cells catalyzed high levels of homologous recombination, whereas very little recombination activity was detected in extracts prepared from normal fibroblasts. Similarly, only extracts derived from immortal cells contained strand-transferase activity as measured by the recently described pairing-on-membrane assay. Mixing experiments indicated that a recombination enhancing factor or factors present in immortal cells, rather than a recombination inhibitor in normal cells, was responsible for the enhanced homologous recombination activity observed using extracts derived from the former.     

Phylogenetic reticulation in subtribe Helianthinae

Incongruence between phylogenetic estimates based on nuclear and chloroplast DNA (cpDNA) markers was used to infer that there have been at least two instances of chloroplast transfer, presumably through wide hybridization, in subtribe Helianthinae. One instance involved Simsia dombeyana, which exhibited a cpDNA restriction site phenotype that was markedly divergent from all of the other species of the genus that were surveyed but that matched the restriction site pattern previously reported for South American species of Viguiera. In contrast, analysis of sequence data from the nuclear ribosomal DNA internal transcribed spacer (ITS) region showed Simsia to be entirely monophyletic and placed samples of S. dombeyana as the sister group to the relatively derived S. foetida, a result concordant with morphological information. A sample of a South American species of Viguiera was placed by ITS sequence data as the sister group to a member of V. subg. Amphilepis, which was consistent with cpDNA restriction site data. Samples of Tithonia formed a single monophyletic clade based on ITS sequence data, whereas they were split between two divergent clades based on cpDNA restriction site analysis. The results suggested that cpDNA transfer has occurred between taxa diverged to the level of morphologically distinct genera, and highlight the need for careful and complete assessment of molecular data as a source of phylogenetic information.     

Antioxidants in peripheral nerve

Oxidative stress and antioxidants have been related in a wide variety of ways with nervous tissue. This review attempts to gather the most relevant information related to a) the antioxidant status in non pathologic nervous tissue; b) the hypothesis and evidence for oxidative stress (considered as the disequilibrium between prooxidants and antioxidants in the cell) as the responsible mechanism of diverse neurological diseases; and c) the correlation between antioxidant alterations and neural function, in different experimental neuropathies. Decreased antioxidant availability has been observed in different neurological disorders in the central nervous system, for example, Parkinson's disease, Alzheimer's disease, epilepsy, amyotrophic lateral sclerosis, cerebral ischaemia, etc. Moreover, the experimental manipulation of the antioxidant defense has led in some cases to interesting experimental models in which electrophysiological alterations are associated with the metabolic modifications induced. In view of the electrophysiological and biochemical effects of some protein kinase C inhibitors on different neural experimental models, special attention is dedicated to the role of this kinase in peripheral nervous tissue. The nervous tissue, central as well as peripheral, has two main special features that are certainly related to its antioxidant metabolism: the lipid-enriched membrane and myelin sheaths, and cellular excitability. The former explains the importance of the glutathione (GSH)-conjugating activity towards 4-hydroxy-nonenal, a biologically active product of lipid peroxidation, present in nervous tissue and in charge of its inactivation. The impairment of the latter by oxidative damage or experimental manipulation of antioxidant metabolism is discussed. Work on different experimental neuropathies from author's laboratory has been primarily used to provide information about the involvement of free radical damage and antioxidants in peripheral nerve metabolic and functional impairment.     

Overall Informativity,OI, in DNA Polymorphisms Revealed by inter-AluPCR: Detection of Genomic Rearrangements

We studied two systems of multilocus markers revealed by PCR using primers directing amplification betweenAlurepeats in a tail-to-tail orientation. Genomic polymorphisms were detected as the presence or absence of the electrophoretic bands representing DNA fragments of a given length. A total of 104 such fragments segregating as Mendelian markers in a panel of eight CEPH families were analyzed by two-point linkage analysis. Fifty-one of these fragments were localized with respect to CEPH markers; they represented 33 loci, 7 of which were multiallelic. Locus-specific oligonucleotides were developed and used as hybridization probes to identify the mapped loci within a complex pattern of inter-AluPCR products. A great proportion of inter-AluPCR polymorphisms represented length variants within amplified DNA segments, while others were presumably due to mutations within the priming sites. To describe the expected number of informative loci per typing experiment we introduced a parameter called overall informativity (OI), which provides a single measure of the multiplex ratio and the informativity of markers contributing to a multilocus system (OIof a single locus is equivalent to its heterozygosity and cannot exceed 0.5 for a biallelic codominant marker). HighOIvalues (5.8 and 11.5) of the two presented systems of inter-AluPCR markers of random chromosomal distribution render them suitable for mapping genomic rearrangements such as genomic deletions in tumoral tissues. This was illustrated by the detection of loss of heterozygosity in the 9q22–qter region in sporadic colon cancer.