Plant regeneration from protoplasts of Dimorphotheca and Rudbeckia

Protoplasts were isolated from leaves, shoots, cotyledons, ray florets and callus cultures of Dimorphotheca aurantiaca (syn. D. sinuata) (Cape Marigold, Star of the Veldt) and Rudbeckia hirta, R. laciniata and R. purpurea; species of ornamental value. For Dimorphotheca, plants were regenerated from protoplasts of all sources apart from the ray floret, whilst for the Rudbeckia species, although protoplast division was induced in most cases, only leaf mesophyll protoplasts of R. hirta c.v. Marmalade gave plants. The establishment of plant regeneration for these ornamental species, from protoplasts, now provides a basis for their somatic hybridisation.Abbreviations BAP 6-benzylaminopurine - IAA indole-3-acetic acid - NAA naphthaleneacetic acid - K kinetin - GA₃ gibberellic acid - MS Murashige and Skoog (1962) - f.wt. fresh weight     

Differential depuration of poliovirus, Escherichia coli, and a coliphage by the common mussel, Mytilus edulis.

The elimination of sewage effluent-associated poliovirus, Escherichia coli, and a 22-nm icosahedral coliphage by the common mussel, Mytilus edulis, was studied. Both laboratory-and commercial-scale recirculating, UV depuration systems were used in this study. In the laboratory system, the logarithms of the poliovirus, E. coli, and coliphage levels were reduced by 1.86, 2.9, and 2.16, respectively, within 52 h of depuration. The relative patterns and rates of elimination of the three organisms suggest that they are eliminated from mussels by different mechanisms during depuration under suitable conditions. Poliovirus was not included in experiments undertaken in the commercial-scale depuration system. The differences in the relative rates and patterns of elimination were maintained for E. coli and coliphage in this system, with the logarithm of the E. coli levels being reduced by 3.18 and the logarithm of the coliphage levels being reduced by 0.87. The results from both depuration systems suggest that E. coli is an inappropriate indicator of the efficiency of virus elimination during depuration. The coliphage used appears to be a more representative indicator. Depuration under stressful conditions appeared to have a negligible affect on poliovirus and coliphage elimination rates from mussels. However, the rate and pattern of E. coli elimination were dramatically affected by these conditions. Therefore, monitoring E. coli counts might prove useful in ensuring that mussels are functioning well during depuration.     

Possible mechanisms for the revival of glutaraldehyde-treated spores of Bacillus subtilis NCTC 8236

Spores of Bacillus subtilis NCTC 8236 were exposed to 2% alkaline glutaraldehyde and subsequently subjected to various treatments in an attempt to revive injured spores. Treatment with alkali (sodium or potassium hydroxide or, to a lesser extent, sodium bicarbonate) proved to be most successful. Some revival was achieved after thermal treatment. No revival was obtained with lysozyme or with various types of coat-removing agents. Experiments designed to distinguish between germination and outgrowth in the revival process established that sodium hydroxide (optimum concentration, 20 mmol/l) added to glutaraldehyde-treated spores increased the potential for germination. In contrast, spores which had been allowed to germinate before exposure to low concentrations of glutaraldehyde and then to sodium hydroxide were inhibited at the outgrowth phase to a much greater extent than germinated spores treated with the dialdehyde without subsequent alkali exposure. The results overall are discussed in terms of the possible mechanism and site of action of glutaraldehyde and the practical implications and significance of its use as a sporicide.     

In vivo brown fat response to hypothermia and norepinephrine in the ovine fetus

The goal of this study was to assess the response of fetal brown fat in vivo to hypothermia and norepinephrine infusion. In 10 unanaesthetized, chronically-prepared fetal sheep (133 +/- 2 days of gestation) cold water was passed through tubing encircling the fetus in utero and plasma glycerol concentration was measured as an indicator of brown fat activity. Following cooling for 60 min, amniotic fluid temperature fell 7.79 degrees C to 31.66 +/- 1.73 degrees C (n = 8, P less than 0.001) and maternal temperature fell 0.63 degree C to 38.63 +/- 0.08 degrees C (n = 9, P less than 0.001). Eight of the fetuses were subjected to a second experiment in which norepinephrine was infused intravenously for 15 min. During infusion fetal arterial temperature fell 0.38 degrees C to 39.05 +/- 0.25 degrees C (n = 7, P less than 0.05). Amniotic fluid temperature (n = 7, NS) and maternal arterial temperature (n = 7, NS) remained constant. Glycerol concentration during the infusion increased from 0.73 to 1.27 mg/dl, a 74% increase over control (n = 8, P less than 0.001). Although clearly detectable, these glycerol responses to hypothermia and norepinephrine stimulation are one-third or less of those achieved after birth, indicating that thermogenesis remains quiescent in the near-term fetal sheep, despite powerful stimuli for activation.     

Simian retrovirus-D serotype 1 (SRV-1) envelope glycoproteins gp70 and gp20: expression in yeast cells and identification of specific antibodies in sera from monkeys that recovered from SRV-1 infection.

The gp70 and transmembrane gp20 envelope proteins of simian retrovirus-D serotype 1 (SRV-1) were expressed in Saccharomyces cerevisiae as fusion proteins with human superoxide dismutase (SOD). Expression of the SOD-gp70 and SOD-gp20 sequences yielded fusion proteins of 52 and 29 kilodaltons, respectively. The yeast-expressed SRV-1 envelope proteins were used in an enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies in the sera of rhesus macaques that recovered from SRV-1. Sera from 47 of 49 such monkeys tested positive for antibodies to the SOD-gp70 fusion protein, while 45 of 49 reacted positively to SOD-gp20. None of 26 SRV-1-nonexposed monkeys tested positive in either ELISA. Monkeys immunized with the recombinant SRV-1 gp20 and gp70 proteins made good ELISA and Western blot (immunoblot) antibodies to whole SRV-1. This antibody was not neutralizing in vitro, however.     

The expression of transthyretin mRNA in the developing rat brain

Specific cDNA and oligonucleotide probes were used to study the appearance of transthyretin mRNA in developing rat brain using Northern gel analysis, cytoplasmic dot hybridization, and in situ hybridization. Transthyretin mRNA in embryonic rat brain was found to be confined to the epithelial layer of the choroid plexus primordia appearing first in the fourth ventricle, followed by appearance in the lateral ventricles, and subsequently in the third ventricle. Transthyretin mRNA was localized in these cells from early stages of neuroepithelium differentiation, showing that it is a sensitive marker for the differentiation of the choroid plexus within the fetal brain.     

Leishmania mexicana: purine-metabolizing enzymes of amastigotes and promastigotes

Cultured promastigote and isolated amastigote forms of Leishmania mexicana mexicana have been surveyed for the presence of enzymes involved in purine metabolism. Quantitative but not qualitative differences between the enzymes of two forms were discovered. There were found to be significant differences between the enzyme content of L. m. mexicana and that reported for L. donovani. Extracts of both parasite forms of L. m. mexicana were found to have higher levels of adenine deaminase (EC 3.5.4.2) and guanine deaminase (EC 3.5.4.3) than adenosine deaminase (EC 3.5.4.4). There appeared to be two distinct nucleosidases (EC 3.2.2.1), one active on nucleosides, the other on deoxynucleosides. Phosphorylase (EC 2.4.2.1) could be detected only in the catabolic direction. Nucleotidases were present, but were more active on 3' (EC 3.1.3.6)- than 5' (EC 3.1.3.5)-nucleotides. Phosphoribosyltransferase (EC 2.4.2.7,.8 and .22) and nucleoside kinase (EC 2.7.1.20) activities were detected in both forms. Nucleotide-interconverting enzymes were found to be present, with IMP dehydrogenase (EC 1.2.1.14) being the most active. Cell fractionation experiments revealed that, in the promastigote, enzyme separation within the parasite may play an important part in regulating cellular purine metabolism.     

Characterization of two molecular weight classes of cadmium binding proteins from the mussel, Mytilus edulis (L.)

Inducible cadmium binding proteins (Cd-BP) in the mussel, Mytilus edulis, were resolved into two molecular weight components by gel permeation chromatography on Sephadex G-75. Each of these two molecular weight components was further resolved into four subcomponents by DEAE ion exchange chromatography. All eight subcomponents bound cadmium and exhibited significant u.v. absorption at 254 and little absorption at 280 nm. Based on amino acid composition analysis two classes of proteins were identified, one having higher cysteine (approximately 25 mole %) and lower serine and glutamic acid contents compared to the other class.     

The nuclear-coded subunits of yeast cytochrome c oxidase. II. The amino acid sequence of subunit VIII and a model for its disposition in the inner mitochondrial membrane

The amino acid sequence of subunit VIII from yeast cytochrome c oxidase is reported. This 47-residue (Mr = 5364) amphiphilic polypeptide has a polar NH2 terminus, a hydrophobic central section, and a dilysine COOH terminus. An analysis of local hydrophobicity and predicted secondary structure along the peptide chain predicts that the hydrophobic central region is likely to be transmembranous. Subunit VIII from yeast cytochrome c oxidase exhibits 40.4% homology to bovine heart cytochrome c oxidase subunit VIIc , at the level of primary structure. Secondary structures and hydrophobic domains predicted from the sequences of both polypeptides are also highly conserved. From the location of hydrophobic domains and the positions of charged amino acid residues we have formulated a topological model for subunit VIII in the inner mitochondrial membrane.