Improvements of the membrane filter method for DNA:rRNA hybridization

We describe and recommend the following improvements of DNA:rRNA membrane filter hybridization methods. One of our aims was to avoid DNA release from filter discs during hybridization.

1. Our hybridization conditions are 2 SSC in aq. dest., with 20% formamide, 50 C, overnight for 16 hr.
2. Duplexing is over in 8–10 hr.
3. Formamide has to be very pure (O.D.≤0.2/cm light path at 270 nm).
4. RNAase treatment: 250 μg/5 ml 2 SSC/filter at 37 C for 1 hr.
5. Our conditions for stepwise thermal denaturation are: 5°C steps from 50C to 90C in 1.5 SSC in 20% formamide.
6. Single-stranded DNA, fixed on membrane filters, and stored in vacuo at 4C, can be used reliably for hybridization for up to 20 months.
7. Concentrated DNA in 0.1 SSC, quick-frozen at ?50 C and stored at ?90 C for up to 2 years can be used for hybridization without much change.
8. A CsCl gradient purification step yields much purer DNA, but increases the release of DNA from filters by about 20%. Filters with 20% more DNA is a compensation.
9. rRNA can be stored for 20 months in SSC or 2 SSC at ?12C without changing the hybridization results.

     

An analysis of amino acid composition of a few oxidoreductases by different methods

1. All methods used to analyze amino acid composition of different enzymes agree well with each other.
2. They lead to a relationship between amino acid composition and function of proteins.
3. Cytochrome oxidase is in a class by itself; copper-containing oxidases are closely related as expected.

     

Effet des androgènes sur l’activité sportive

Androgenic hormones seem to be of beneficial effects on sports performance:

- they increase motivation, will, aggressiveness, resistance to the stress and to the fatigue, leading to an increase of the training quantity,

- they increase bone mineralization and probably mechanical resistance,

- they stimulate the bone marrow and so, with the erythropoietin, the erythropoiesis,

- they increase the tendancy to hyperglycemia, but with a decrease of the tolerance to the glucose,

- they stimulate the fatty acids mobilization from the adipose tissue, for their utilization in the muscle during the exercise,

- they participate, for the trained sportmen, to a better gestion of the muscle glycogen storage: their utilization during exercise is decreased,

- they increase the lean body mass, with an increase of the protein synthesis and a decrease of the protein catabolism, leading also to an increase of the muscle force under training. There is no beneficial effect upon the tendons,

- they have an immunomodulation action.

     

1,2,3-Thiadiazoles as a Convenient Source for the Study of Molecular Rearrangements,Single Bond/No Bond Resonance and Dendrimer Synthesis

1,2,3-Thiadiazoles are five-membered heterocycles which are readily available. We have used these interesting compounds in three different areas of research:

• ? Rearrangements of 1,2,3-thiadiazoles leading to other heterocycles, such as 1,2,3-triazoles and 1,2,3,4-thiatriazoles. Isomeric 1,2,3-thiadiazoles (a ring-degenerate rearrangement) could also be obtained.
• ? 1,2,3-Thiadiazolium salts as synthons for 6aλ⁴-thiapentalenes showing single bond/no bond resonance, and/or mesoionic compounds.
• ? The base-induced cleavage of 1,2,3-thiadiazoles giving the reactive alkynethiolates, which were used in the synthesis of dendrimers.

     

The influence of EDTA on the composition of alginate synthesized by Azotobacter vinelandii

1. During an investigation of the physiology of Azotobacter vinelandii with particular reference to polysaccharide formation, a suitable medium which was precipitate-free was developed by adding EDTA at a concentration of 50 mg/l to a basal medium containing one of eight different carbohydrates as sole carbon source.
2. Acetylated alginate was always produced by the organism when cultured under defined conditions, regardless of the carbohydrate source incorporated in the basal medium.
3. When EDTA was added to the medium, the bacteria produced acetylated polyuronides with a preponderance of mannuronic acid residues.
4. A comparison of the infrared spectra of the alginate produced by Azotobacter vinelandii and the affect of EDTA upon the mannuronic acid/guluronic acid ratios of the alginate are reported.

     

Protease and Amylase in the digestive tract ofBarbus paludinosus

1. Protease and amylase activity in the digestive system ofBarbus paludinosus Peters (Pisces, Cyprinidae) has been investigated.
2. Chromatographic analysis showed seven amino acids to be present in both the anterior and posterior intestine. Only leucine, phenylalanine, valine, glycine and aspartic acid were positively identified.
3. In the anterior intestine chromatography revealed two sugars, but only one in the posterior intestine which was identified as glucose.
4. The pH of the intestinal fluid was found to be 5.8 and 7.8 for the fore and hind gut respectively, This correlates well with the enzyme pH optima found in in vitro experiments.
5. Protease and amylase activity was found throughout the digestive tract. Maximum proteolytic activity being present in the anterior intestine. Amylase activity is similar in both regions of the gut.
6. Correlation between the digestive enzymes and the fishes diet is briefly discussed.

     

Studies on the origin of the tip potential of glass microelectrode

1. Tip potential (TP) of glass microelectrodes filled with 3 M KCl increased remarkably with the increase in the storage period in 3 M KCl solution at 37? C, while the electrode resistances decreased gradually.
2. The electrical conductivity through the thin glass wall near the tip was found to increase in parallel with the TP increase.
3. The e.m.f. across the thin glass wall in the tip region was directly measured. This seems to contribute to the TP generation of the microelectrode when the conductivity of the glass wall is significantly high in the tip region.
4. Effects of the acid treatment of glass employed and the acidification of fillant electrolyte solution suggested that fixed negative charges on the glass wall play a fundamental role in the TP formation.
5. Based on these experimental results, it was concluded that not only the diffusion potential through the tip pore but also the interfacial potential through the thin glass wall near the tip contributes to the TP generation, and the contribution of the latter increases with a long exposure period of the electrodes to electrolyte solution.
6. In this connection, technical problems related to reduction of the tip potential were also discussed.

     

Charkterisierung eines phagenähnlichen Partikels aus Zellen von Nitrobacter

1. Phage-like particles Nb₁ isolated from cells of Nitrobacter agilis were characterized after freeze etching and after treatment by fixation agents.
2. Ethanol-acetic acid fixed particles can be digested by the proteolytic enzyme papain.
3. Ethanol-acetic acid fixed particles show a loss in mass and volume after treatment with DNase. Under the same conditions RNase has no influence.
4. The chemical composition of the phage-like particle Nb₁ is discussed.

     

Succession in Zoo- and phytoplankton in a sewage stabilization pond

The investigations pertaining to zoo- and phytoplankton succession during prestabilization and stabilization of domestic waste water in a facultative pond operating at 1.22 meters depth with an average BOD loading of 336.3 kg per hectare per day and a retention period of 10 days revealed the following:

1. A photosynthetic sulphur bacterium Thiocystis violacea has been recorded during the second and third day along with a considerable population of ciliate protozoa and algae.
2. Chlorophytes were followed by euglenoids in the ecological succession and the latter became predominant after the pond stabilized.
3. Of the twelve protozoan species Halteria grandinella, Bursaria truncatella and Perispira ovum were associated with peaks of algae and vigorous photosynthesis in the stabilized pond water.
4. There ia a gradual improvement in the quality of the waste water with respect to reduction in bacteria and biochemical oxygen demand and enhancement in changes in physico-chemical factors till a relatively stable community is evolved.

     

Biogenesis of mitochondria 20 the effects of altered membrane lipid composition on mitochondrial oxidative phosphorylation inSaccharomyces cerevisiae

1. The lipid composition of mitochondria isolated from a fatty acid desaturase mutant ofSaccharomyces cerevisiae may be extensively manipulated by growing the organism on defined supplements of unsaturated fatty acid (UFA).
2. The fatty acid composition of the mitochondrial lipids closely follows that of the whole cells from which the mitochondria are isolated. UFA-depleted mitochondria contain normal levels of sterols, neutral lipids and total phospholipids, but have much lower levels of phosphatidyl inositides.
3. UFA-depleted mitochondria possess a full complement of cytochromes, oxidase both NAD-linked and flavoprotein-linked substrates at normal rates, and have levels of succinate and malate dehydrogenases similar to those of UFA-supplemented mitochondria. However, UFA-depletion has a marked effect on the ability of cytochromec to reactivate the NADH oxidase activity of cytochromec-depleted mitochondria.
4. The efficiency of oxidative phosphorylation decreases progressively with the UFA content of the mitochondria, and oxidative phosphorylation is completely lost in mitochondria containing approximately 20% UFA.
5. The incorporation of UFA into the lipids of UFA-depleted mitochondriain vivo results in a recoupling of oxidative phosphorylation. Recoupling is insensitive to both chloramphenicol and cycloheximide, indicating that all the proteins necessary for oxidative phosphorylation are present in UFA-depleted mitochondria, and that the less of oxidative phosphorylation is a purely lipid lesion.
6. ATPase activity is apparently unaffected by UFA-depletion, but³²P_i-ATP exchange activity is lost in mitochondria which have been extensively depleted in UFA.
7. Valinomycin stimulates the respiration of UFA-supplemented mitochondria in media containing potassium, but has no effect on the respiration of UFA-depleted mitochondria, suggesting that active transport of potassium is lost as a result of UFA-depletion.

     

Context-dependent stuttering

Bei einer Untersuchung des Stotterns in der deutschen Umgangssprache machten wir folgende Beobachtungen:

1. Das gestotterte Phonem wird häufig von einem identischen Phonem begleitet (definiert als das ?induzierende Phonem“) welches sowohl vor wie nach dem gestotterten Phonem auftreten kann.
2. Gewöhnlich folgte das induzierende Phonem dem gestotterten Phonem.
3. Der Abstand zwischen induzierendem und gestottertem Phonem war geringer als bei Zufälligkeit zu erwarten.
4. Induzierende und gestotterte Phoneme befanden sich gewöhnlich in identischen Silbenpositionen.
5. Gestotterte Phoneme traten in der Regel bei betonten Silben auf.

Um diese Beobachtungen zu erklären erschienen uns drei Annahmen erforderlich:

1. Sprach-Output ist hierarchisch bestimmt. Silben und Phoneme sind Glieder in dieser Hierarchie.
2. Unterschwellige Erregbarkeit ist in dieser Hierarchie stärker bei betonten als bei unbetonten motorischen Programmen.
3. Ähnliche Programme (sowohl auf Silbenals auch Phonemniveau) inhibieren einander.

Diese Annahme gibt zugleich eine mögliche Erklärung für Blockierung und Längung — Phänomene, die ebenfalls in der Sprache von Stotterern auftreten. Unsere Beobachtungen bieten also eine mögliche Lösung für das Rätsel des Stotterns.     

EPR characterization of oxide supported transition metal ions: Relevance to catalysis

EPR spectroscopy is a powerful tool to identify at a molecular level, the different steps of catalyst preparation, and of catalytic reactions:

1. Deposition of paramagnetic transition metal ions onto a support is monitored, and the coordination sphere of the metallic center is characterized by EPR.
2. The catalyst is also characterized after activation (thermal oxidation or reduction):

- the distribution among the different sites in zeolites can be determined;

- the dispersion of the active phase may be appreciated;

- the unsaturation degree of the active site may be evaluated using probe molecules such as water or¹³C enriched carbon monoxide.

1. The catalytic mechanisms can be investigated by studying the elementary steps of the catalytic reaction, as illustrated for methanol oxidation over Mo/SiO₂ catalysts whose EPR results have extended the reaction mechanism proposed on the basis of kinetic data. In addition, reaction intermediates may be isolated inquasi-in situ conditions as in the case of olefin oligomerization catalyzed by Ni/SiO₂ systems.

     

Life cycle assessment study on resilient floor coverings

Fourteen European producers of resilient floor coverings examined thirty-two objects of their products in a Life Cycle Assessment Study. The product groups were PVC, cushioned PVC, polyolefin, rubber and linoleum, with one reference example from textile and parquet. Important results include the following:

? There is no material specific ranking for ‘best’ or ‘worst’ environmental performance. Differences within the material groups - depending on the individual formula - are larger than between the groups.

? The introduction of material-specific recycling for used floorings as well as the use of recycled material in the flooring production could reduce environmental loads significantly.

? The premature change of a flooring by the user may induce a major influence on the environmental performance of a flooring.

     

Les responsabilités du biologistes

The responsabilities of the professionals are numerous and various. As an exemple, we discuss the french law (29 july 1994) about the practice of reproductive biology. There are three levels of responsabilities:

? civil liability concern the damage to somebody, the means, information, confidentiality.

? penal code concern the integrity of somebody, the safety of the staff and of the patients.

? administrative.

All the professionals must be aware of the legislation and the penalties they might incur.     

Effect of external factors on phototaxis of Chlamydomonas reinhardtii

1. A fully automated phototaxis monitoring device is described for measuring photo-topatactic responses of flagellated organisms.
2. Photokinesis can be demonstrated in Chlamydomonas cells only after a dark period of about 72 hrs.
3. Pre-darkening of a few hours duration raises the phototactic disposition, whereas pre-illumination has no significant effect.
4. Circadian rhythms can be initiated by only one period of darkness or lower light intensity, whereas a period of higher intensity does not induce rhythms. The period length of the circadian rhythms is about 24 hrs.

     

The use of a continuous assay system to show the stimulation of phosphoenolpyruvate production in intact mitochondria by valinomycin and by Mn2+

1. A method for the direct recording of the PEP efflux from isolated mitochondria is described.
2. This method has been used to show the stimulation of PEP efflux by externally added Mn⁺⁺ ions.
3. Valinomycin, uncoupler and oleate were also shown to stimulate PEP efflux.
4. Valinomycin caused an increase in the internal concentration of both PEP and citrate.
5. The results indicate that the major pathway of PEP synthesis in isolated mitochondria is via PEP carboxykinase and the results do not call for an unknown pathway of metabolism.
6. Two interactions between PEP and citrate are described; competition for the mitochondrial interior and the stimulation of PEP production by citrate.

     

Metabolism of the anaerobic formation of succinic acid bySaccharomyces cerevisiae

1. Succinic acid is formed in amounts of 0.2–1.7 g/l by fermenting yeasts of the genusSaccharomyces during the exponential growth phase. No differences were observed between the various species, respiratory deficient mutants and wild type strains.
2. At low glucose concentrations the formation of succinic acid depended on the amount of sugar fermented. However, the nitrogen source was found to be of greater importance than the carbon source.
3. Of all nitrogen sources, glutamate yielded the highest amounts of succinic acid. Glutamate led to an oxidative and aspartate to a reductive formation of succinic acid.
4. A reductive formation of succinic acid by the citric acid cycle enzymes was observed with malate. This was partially inhibited by malonate. No evidence was obtained that the glyoxylate cycle is involved in succinic acid formation by yeasts.
5. Anaerobically grown cells ofSaccharomyces cerevisiae contained α-ketoglutarate dehydrogenase. Its activity was found in the 175000 x g sediment after fractionated centrifugation. The specific activity increased 6-fold after growth on glutamate as compared with cells grown on ammonium sulfate.
6. The specific activities of malate dehydrogenase, fumarase, succinate dehydrogenase, succinylcoenzymeA synthetase, α-ketoglutarate dehydrogenase and glutamate dehydrogenase (nicotinamide adenine dinucleotide dependent) were determined in yeast cells grown on glutamate or ammonium sulfate. Similar results were obtained with a wild type strain and a respiratory deficient mutant. The latter did not contain succinate dehydrogenase.
7. In fermenting yeasts succinic acid is mainly formed from glutamate by oxidation.

     

A study on the rate of water transport of the clam katelysia opima in relation to environmental conditions

1. The neutral red technique was employed to study the rate of filtration in Katelysia opima.
2. The weight specific water filtration was found to be greater for younger clams compared to the older ones.
3. The rate of water filtration increased with decreasing salinity.
4. Water filtration was found to increase as temperature increased, reaching a maximum at 35°C. but then sharply decreasing at 39°C.
5. Light had no significant effect on the rate of filtration.
6. Suspended matter was found to affect the rate of water filtration.
7. The rate of filtration was low at high pH and high in low pH.
8. The rate of water filtration was found to be faster during high tide than during low tide.
9. The presence of the parasitic crab, Pennotheris sp., in the mantle cavity of clams had a marked effect on the particle filtration.
10. Accidental cut of the siphon tips had no effect on the rate of filtration.