Determination of Gardnerella vaginalis Genome Size by Pulsed-Field Gel Electrophoresis

The chromosomal DNA of four strains of Gardnerella vaginaliswere digested with rare cutting restriction enzymes and analyzedby pulsed-field gel electrophoresis (PFGE). The four strainsstudied were two clinical isolates (GVP 004 & GVP 007) andtwo American Type Culture Collection strains (ATCC 14018 &ATCC 14019). The restriction enzyme SfiI generated two DNA fragmentsof about 0.6 Mb and 1.1 Mb in all four strains giving a G. vaginalisgenome size of about 1.7 Mb. A similar genome size was calculatedutilizing two more GC-rich sequence specific restriction endonucleases,NotI and AscI. When digested with AscI, the chromosomal DNAof all four strains gave rise to 11 to 12 DNA fragments rangingbetween 0.01 Mb to 0.43 Mb. DNA from the two clinical isolateswere digested by NotI (yielding 7 to 9 fragments), while theDNA from the two ATCC strains were resistant to NotI digestion.In contrast to the clinical isolates, DNA from the two ATCCstrains gave an identical profile for all restriction endonucleasestested. From double digestion experiments, the two SfiI sitescould be localized on two AscI fragments. From these PFGE studies,it is concluded that the G. vaginalis genome is a circular DNAthat ranges between 1.67 Mb and 1.72 Mb in size.     

Application of continuous zone electrophoresis to preparative separation of proteins

A comprehensive study of the application of continuous zone electrophoresis to preparative separation of proteins in free solution is presented. First, the influence of electric field strength, buffer residence time in the chamber, sample flow rate, and sample concentration on separation resolution and throughput were studied. Using multiple injections of sample into the electrophoresis chamber, a throughput of 500 mg protein/h was achieved for partially purified model proteins. Experiments on Escherichia coli crude extracts yielded a fivefold purification of beta-galactosidase along with a simultaneous separation of proteins from cell debris in a single step. Experiments correlating the electrophoretic mobility in continuous electrophoresis with the elution behavior in ion-exchange chromatography were performed on more than a dozen proteins which conclusively showed that separation of proteins in continuous zone electrophoresis is governed by net surface charge. Based on these results, the fraction numbers in which the proteins eluted could be correctly predicted. Proteins and enzymes with differences >0.5 M elution molarities in ion-exchange chromatography were separated by continuous zone electrophoresis on a preparative scale (mg/h or g/h) with >90% recovery. This corresponds to a preparative scale separation of proteins and enzymes which differ in apparent electrophoretic mobility by only 0.70 x 10(-5) cm(2)/V . s. (c) 1993 John Wiley & Sons, Inc.     

Phytochemical Characterization of Cannabis sativa L. Roots from Northeastern Brazil

This article describes the phytochemical study of Cannabis sativa roots from northeastern Brazil. The dried plant material was pulverized and subjected to exhaustive maceration with ethanol at room temperature, obtaining the crude ethanolic extract (Cs-EEBR). The volatile compounds were analyzed by gas chromatography coupled with mass spectrometry (GC/MS), which allowed to identify 22 compounds by comparing the linear retention index (LRI), the similarity index (SI) and the fragmentation pattern of the constituents with the literature. By this technique the major compounds identified were: friedelan-3-one and β-sitosterol. In addition, two fractions were obtained from Cs-EEBR by classical column chromatography and preparative thin layer chromatography. These fractions were analyzed by NMR and IR and together with the mass spectrometry data allowed to identify the compounds: epifriedelanol, friedelan-3-one, β-sitosterol and stigmasterol. The study contributed to the phytochemical knowledge of Cannabis sativa, specifically the roots, as there are few reports on the chemical constituents of this part of the plant.     

Possible causes of the physiological decline in soybean nitrogen fixation in the presence of nitrate

Nodulated soybean plants (Glycine max (L.) Merr. cv. Clarke)were supplied with 10 mol m^-3 nitrate at the vegetative stage.This treatment caused a rapid decline in nitrogen fixation (acetylenereduction) activity and a consequent decline in ureides in thexylem sap. However, there was virtually no effect on the nitrogenasecomplex, according to Western blots against components 1 and2. The effect on nitrogen fixation was matched by a decreasein nitrogenase-linked respiration and increases in nodule oxygendiffusion resistance and the carbon cost of nitrogen fixation.The addition of nitrate had little effect on protein contentfrom either nodule plant or bacteroid fractions. Activitiesof nitrate reductase (NR) and nitrite reductase (NiR) from eitherthe plant fraction or the bacteroids were affected in differentways during 8 d of supply. Nodule plant NR and bacteroid NiR were not affected. However,nodule plant NiR increased 5-fold within 2 d of supplying Bacteroid NR only increased after6 d. These results could be interpreted in terms of a restrictednitrate access into the infected region of nodules. However,denitrification was detected within 2 d of nitrate supply insoybean nodules. The results are discussed in relation to possiblecauses of the nitrate-induced decline in nitrogenase activity. Key words: Glycine max, nitrate, nitrogen fixation, nodules     

Additional evidence implicating Triticum searsii as the B-genome donor to wheat

In vitro DNA:DNA hybridizations and hydroxyapatite thermal-elution chromatography were employed to identify the diploid wheat species ancestral to the B genome of Triticum turgidum. ³H-T. turgidum DNA was hybridized to the unlabeled DNAs of T. urartu, T. speltoides, T. sharonensis, T. bicorne, T. longissimum, and T. searsii. ³H-Labeled DNAs of T. monococcum and a synthetic tetraploid AADD were hybridized with unlabeled DNAs of T. urartu and T. searsii to determine the relationship of the A genome of polyploid wheat and T. urartu. The heteroduplex thermal stabilities indicated that T. searsii was most closely related to the B genome of T. turgidum (AB) and that the genome of T. urartu and the A genome have a great deal of base-sequence homology. Thus, it appears that T. searsii is the B-genome donor to polyploid wheat or a major chromosome donor if the B genome is polyphyletic in origin.Published with the approval of the Director of The West Virginia Agricultural Experiment Station as Scientific Paper No. 1837.     

Intestinal oxalate uptake in castrated male and female rats: evidence for altered brush border membrane composition

The intestinal uptake rate of oxalate (mumoles/h/g tissue wt.) in castrated male (CM) rats, CM rats administered estradiol, and female (F) rats was 1.8, 1.4 and 1.3 times higher than that of male rats, whereas castrated female (CF) rats and CF rats administered testosterone absorbed oxalate at a rate similar to F rats, thereby, suggesting that gonadectomy affected intestinal uptake of oxalate only in male rats The intestinal oxalate uptake rate in all the groups increased linearly with increasing oxalate concentration (0.1- 6.0 mM). Chemical composition of brush border membrane showed significant changes in the sialic acid, phospholipid and cholesterol content following castration, which may lead to ultrastructural changes in the membrane thereby, increasing the absorption of oxalate.     

Possible mitochondrial involvement in mechanism of cytoplasmic male sterility in maize (Zea mays L.)

The mechanism of cytoplasmic male sterility was investigated in maize by isolating mitochondria from seedlings and various anther stages and analyzing cytochrome oxidase and succinic dehydrogenase biochemically and electrophoretically. Sterile anthers exhibited a lack of biochemical activity and fewer isozymatic bands for cytochrome oxidase. No apparent differences were detected biochemically or electrophoretically between fertile and sterile anthers for succinate dehydrogenase.     

Pretranslational regulation of alpha 2u-globulin in rat liver by growth hormone

Hypophysectomy is known to cause complete suppression of the hepatic synthesis alpha 2u-globulin. The effect of hypophysectomy on the synthesis of alpha 2u-globulin can be reversed by multiple hormone treatment. The role of pituitary growth hormone in the multihormonal regulation of alpha 2u-globulin in rat liver was examined in the hypophysectomized male rats with and without growth hormone supplementation. Daily treatment of hypophysectomized rats with 5 alpha-dihydrotestosterone, corticosterone, thyroxine, and growth hormone for 8 days caused about 80% recovery in the hepatic content of alpha 2u-globulin and its corresponding mRNA as determined by radioimmunoassay, in vitro translation, and liquid hybridization with a cloned cDNA probe. However, omission of growth hormone from the treatment regimen failed to raise hepatic alpha 2u-globulin and its mRNA to more than 5% of the normal control. The possible effect of growth hormone on the translation of the mRNA for alpha 2u-globulin was examined with cultured hepatocytes derived from growth hormone-deficient rats. Culture of these cells in the presence of growth hormone for 24 h did not turn on the synthesis of alpha 2u-globulin. These results indicate that growth hormone regulates the synthesis of alpha 2u-globulin by acting at a step antecedent to mRNA translation.     

Genetics of seed-coat colour in Citrullus lanatus (Thunb.) Mansf.

Summary Intervarietal crosses in watermelon, Citrullus lanatus (Thunb.) Mansf., involving six parents with black (J18-1 and J 75), brown (J56-1 and N.H. Midget), red (Bykovski-199) or light cream (Red Nectar) seed-coat colour were made. Parents, F₁, F₂ and backcross populations were evaluated for their phenotypic expressions with regard to the seed-coat colours involved. Black colour was monogenically dominant over brown light cream and red colour of seed-coat separately or independently. Red colour was dominant over light cream colour of seed-coat by a single pair of genes. The light cream colour was recessive to the brown seed-coat colour of watermelon where a single pair of genes was involved.