Isolation of methotrexate-resistant cell lines in Petunia hybrida upon stepwise selection procedure

Summary Cell suspensions of Petunia hybrida were subjected to a selection procedure in which the concentration of the selective agent, methotrexate (MTX), was gradually elevated. In mammalian cells, this procedure frequently results in MTX-resistant mutants due to amplification of the gene coding for dihydrofolate reductase (DHFR), the target protein of MTX.Five suspension lines were isolated, with degrees of resistance ranging from 10 to 500 M MTX (in wild type the LD_99.9 is 0.2 M). MTX^R phenotypes were unstable, as manifested by the loss of resistance upon prolonged growth in the absence of drug. All of the mutants also exhibited high values of MTX-binding protein (60- to 400-fold higher than that of the wild type), which declined to intermediate values upon MTX withdrawal. Finally, cellular extracts from all of the mutants also showed high specific staining of DHFR-activity in gels.The results suggest that the resistance of MTX in these plant cell-lines is mediated by the elevation of the amounts of DHFR, probably as a consequence of gene amplification.     

Microspatial heterogeneity in the distribution of ciliates in a small pond

Five transects of contiguous samples from the surface of a small pond and one transect from its bottom were collected in order to quantify microspatial heterogeneity in the distribution of ciliated protozoa. Examination of the frequency-abundance relations for these transects suggests that they can be approximated by negative binomial distributions with a commonk of 1.87. Contagiousness or crowding increases with population density.Mean patch size and mean interpatch distance were measured for 4 transects as 1.5 to 2 cm and 3 to 4 cm, respectively. This heterogeneity is suggested to arise from behavioral aggregation about discrete food sources and be very ephemeral.Blocking of adjacent contiguous samples was used to investigate the effect of sample size on the apparent correlation between the numbers of pairs of taxa. In all cases examined, taxa were relatively independent in their distribution at small sample sizes and became more negatively or positively associated as samples were combined. This may reflect that the small scale patches are essentially monospecific.     

Contribution to the structural characterization of eucaryotic PSI reaction centre—I. Critical analysis of the polypeptidic composition of different P700 enriched fractions

In thylakoid membranes, several peptides of high MW are present which may interfere with the study of CP1's components. Modifying Cleveland's technique [7] for limited proteolysis, we have characterized the polypeptides found in the 60 kD region. Some may result from incomplete washing of the CF1 while others come from the CP1; indeed, this chlorophyll protein complex, which has a higher MW (above 100 kD), very often undergoes a dissociation into smaller components of about 60 KD MW.Analysis of the protein content of different preparations commonly used to obtain PSI reaction centre enriched fractions has been performed. The and subunits of CF1 are among the main contaminants of most of these preparations. A further purification step is described which can be applied to all these preparations, but numerous peptides are still present in the active fractions. It is most unlikely that all these polypeptides are required for the primary photochemical event, and this emphasizes the necessity to find a new simple method to purify PSI reaction centres.     

Amylolytic enzymes from Aspergillus hennebergi (A. niger Group): Purification and characterization of amylases from solid and liquid cultures

Two glucoamylases (I and II) were produced during solid-state culture of Aspergillus hennebergi (A. niger group) on cassava meal, whereas one glucoamylase and one alpha-amylase were synthesized by the mould in liquid culture. These glucoamylases were acidic proteins with thermotolerant activities. Glucoamylase I was not a glycoprotein, but glucoamylase II and the glucoamylase from liquid cultures contained 15% of sugars. The alpha-amylase was significantly less thermotolerant and of smaller molecular weight. The influence of culture conditions on the production of different amylases by the same Aspergillus strain on the same substrate is discussed.     

ZINC ADSORPTION AND TRANSPORT BY CHLAMYDOMONAS VARUIABILIS AND SCENEDESMUS SUBSPICATUS (CHLOROPHYCEAE) GROWN IN SEMICONTINUOUS CULTURE1

The amount of zinc adsorbed onto the cell surface of the unicellular green algae Scenedesmus subspicatus Hodat and Chlamydomonas variabilis Dangeard was operationally defined by extraction with EDTA; it was a function of the concentration of free ionic zinc remaining in the growth medium, rather than that of the total (free plus complexed) zinc concentration, and could be described by Langmuir isotherms. Conditional adsorption equilibrium constants for zinc were 0.123 and 0.039 L ·μmol^?1 for S. subspicatus and C. variabilis, respectively. A portion of the zinc adsorbed onto C. variabilis was released into solution after 1 h of contact with the metal, providing a possible tolerance mechanism for this alga; the division rate of C. variabilis was not altered by up to 12 μmol Zn²⁺· L^?1, although the cell yield obtained during the stationary phase was significantly decreased. The amount of transported or cellular zinc, for both algal species, was operationally defined as the zinc remaining with the cell after EDTA-extraction; it was a linear function of the free ionic zinc concentration remaining in solution, suggesting that the zinc transported into the cell was not derived from the total adsorbed fraction, although the latter may contain some zinc originating from specific sites leading to zinc transport.     

Photomodulation of growth and specific changes in fatty acid amounts in internodes of green Vigna sinensis L.

First internode growth of green Vigna sinensis L. can be widely modified by light or dark treatments. In all the treatments used there is a good correlation between the internode growth and the rate of C_18-1 accumulation. None of the other fatty acids show such a correlation.Abbreviations C_16-0 palmitoic acid - C₁₇ heptadecanoic acid - C_18-0 stearic acid - C_18-1 octadecenoic acid - C_18-2 linoleic acid - C_18-3 linolenic acid - D darkness - DW dry weight - FR far-red light - FW fresh weight - P_FR phytochrome in the FR absorbing form - R red light - W white light     

Ans fluorescence and electrokinetic potential of activated lymphocytes by con A or LPS in vivo

The $\text{in vivo}$ effect of Concanavalin A (Con A), or bacterial lipopolysaccharide (LPS) on mouse spleen cell populations was investigated. The membrane fluorescence changes of activated splenic lymphocytes were studied during two weeks after the injection of polyclonal immunogens. Experiments were performed with the hydrophobic fluorescent probe: 1-anilino-8-naphthalene sulphonate (ANS). The kinetic studies further indicated that the course of fluorescence changes may considerably vary depending on both immunogens. These fluorescence intensity changes would be in direct relation to the electrokinetic surface potential changes of activated lymphocytes, as assessed by the electrophoretic mobility analysis. By comparison with the inverse relationship observed in our previous study, it would be concluded that the relation (direct or inverse) between ANS fluorescence and electrokinetic potential depends on the net electrical charge of the antigens used.