Longitudinal study of a heteroplasmic 3460 Leber hereditary optic neuropathy family by multiplexed primer-extension analysis and nucleotide sequencing.

Nucleotide-sequencing and multiplexed primer-extension assays have been used to quantitate the mutant-allele frequency in 14 maternal relatives, spanning three generations, from a family that is heteroplasmic for the primary Leber hereditary optic neuropathy (LHON) mutation at nucleotide 3460 of the mitochondrial genome. There was excellent agreement between the values that were obtained with the two different methods. The longitudinal study shows that the mutant-allele frequency was constant within individual family members over a sampling period of 3.5 years. Second, although there was an overall increase in the mutant-allele frequency in successive generations, segregation in the direction of the mutant allele was not invariant, and there was one instance in which there was a significant decrease in the frequency from parent to offspring. From these two sets of results, and from previous studies of heteroplasmic LHON families, we conclude that there is no evidence for a marked selective pressure that determines the replication, segregation, or transmission of primary LHON mutations to white blood cells and platelets. Instead, the mtDNA molecules are most likely to replicate and segregate under conditions of random drift at the cellular level. Finally, the pattern of transmission in this maternal lineage is compatible with a developmental bottleneck model in which the number of mitochondrial units of segregation in the female germ line is relatively small in relation to the number of mtDNA molecules within a cell. However, this is not an invariant pattern for humans, and simple models of mitochondrial gene transmission are inappropriate at the present time.     

Optical Fiber-Based In Situ Spectroscopy of Pigmented Single Colonies

We have adapted a commercially available fiber-optic spectroradiometer with diode array detection to record reflection and absorption spectra from single, 1-mm-diameter bacterial colonies. A careful assessment of the performance of the spectroradiometer for this application is reported. In a model study employing colonies from various phototrophic bacteria, we show that the reflectance spectra are reliable within the range of 450 to 820 nm, whereas the transmission spectra yield accurate peak intensities and absorption maxima from 400 to 900 nm. For screening of populations of about 10(sup4) colonies, fiber-optic transmission spectroscopy provides an attractive and inexpensive alternative to present techniques based on charge-coupled device imaging technology.     

Immunoblotting identifies and antigen recognized by anti gp63 in the immune complexes of Indian kala-azar patient sera

In SDS-PAGE the immune complexes (IC) of kala-azar patient sera showed intense bands at 55 kDa and 20 kDa corresponding to heavy and light chains of immunoglobulins. In immunoblot experiment, kala-azar and normal IC after treatment with patient sera showed multiple bands of which the band at 55 kDa was most prominent in kala-azar IC. It is known that in kala-azar sera antihuman IgG is present, so the heavy band at 55 kDa region may be due to higher amount of IgG and/or other antigen(s) present at that region. Immunoblot experiments of kala-azar IC with anti gp63 also developed a major band at 55 kDa. It suggests that the antigen (55 kDa) and gp63 have common antigenic epitope (s). Normal IC did not react with anti gp63 indicating absence of this antigen in normal IC. Antigenic similarity between the IC antigen (55 kDa) and gp63 indicated that the former antigen may have been processed from gp63. In summary, identification of a parasite antigen (55 kDa) in IC of kala-azar patients sera may be useful in developing a serodiagnostic assay for visceral leishmaniasis. (Mol Cell Biochem130: 11–17, 1994)Abbreviations IC Immune Complexes - PEG Polyethylene Glycol (Mol wt 8000) - PBS Phosphate Buffer Saline - VL Visceral Leishmaniasis - AVL American Visceral Leishmaniasis - IgG Immunoglobulin G - TBS Tris Buffer Saline - SDS-PAGE Sodium Dodecyl Sulphate Polyacrylamide Gel Electrophoresis - gp63 A leishmanial surface glycoprotein of molecular mass 63,000 - TEMED N,N,N,N-Tetramethylethylenediamine     

Molecular cloning and sequencing of an operon, carRS of Azospirillum brasilense, that codes for a novel two-component regulatory system: demonstration of a positive regulatory role of carR for global control of carbohydrate catabolism.

A pleiotropic carbohydrate mutant, CR17, of Azospirillum brasilense RG (wild type) that assimilates C4 dicarboxylates (succinate and malate) but not carbohydrate (fructose, arabinose, galactose, glycerol, and gluconate) as C sources for growth was used to identify the car (carbohydrate regulation) locus by complementation analysis. The 2.8-kb genomic fragment that complemented the Car- defect of CR17 and overlapped the fru operon (S. Chattopadhyay, A. Mukherjee, and S. Ghosh, J. Bacteriol. 175:3240-3243, 1993) has now been completely sequenced. The sequence contains an operon, carRS, coding for two proteins, CARR and CARS, having 236 and 352 amino acid residues, respectively. The 3'-flanking region of the carRS operon showed sequence homology with the 5' terminus of the fruB gene of a related bacterium, Rhodobacter capsulatus. A complementation study with carRS deletion clones showed that only the carR+ gene was required to complement the Car- defect of CR17, signifying that the carbohydrate pleiotropy was due to a lesion within this gene. Although the 2.8-kb DNA containing the carRS operon when introduced by conjugation into CR17 also complemented the Car- defect, the complemented transconjugant was unable to utilize succinate as a C source. The reason for this is not clear. A sequence analysis of the two protein products strongly suggests that the protein pair may constitute a novel two-component regulatory system for global expression of carbohydrate catabolic pathways in A. brasilense.     

Effects of deletions in the carboxy-terminal hydrophobic region of herpes simplex virus glycoprotein gB on intracellular transport and membrane anchoring.

The gB glycoprotein of herpes simplex virus type 1 is involved in viral entry and fusion and contains a predicted membrane-anchoring sequence of 69 hydrophobic amino acids, which can span the membrane three times, near the carboxy terminus. To define the membrane-anchoring sequence and the role of this hydrophobic stretch, we have constructed deletion mutants of gB-1, lacking one, two, or three predicted membrane-spanning segments within the 69 amino acids. Expression of the wild-type and mutant glycoproteins in COS-1 cells show that mutant glycoproteins lacking segment 3 (amino acids 774 to 795 of the gB-1 protein) were secreted from the cells. Protease digestion and alkaline extraction of microsomes containing labeled mutant proteins further showed that segment 3 was sufficient for stable membrane anchoring of the glycoproteins, indicating that this segment may specify the transmembrane domain of the gB glycoprotein. Also, the mutant glycoproteins containing segment 3 were localized in the nuclear envelop, which is the site of virus budding. Deletion of any of the hydrophobic segments, however, affected the intracellular transport and processing of the mutant glycoproteins. The mutant glycoproteins, although localized in the nuclear envelope, failed to complement the gB-null virus (K082). These results suggest that the carboxy-terminal hydrophobic region contains essential structural determinants of the functional gB glycoprotein.     

Design and synthesis of polyacrylamide-based oligonucleotide supports for use in nucleic acid diagnostics.

Polyacrylamide supports, in a range of pore sizes, were investigated as nucleic acid affinity matrices for the detection of target DNA or RNA sequences using a sandwich hybridization format. Bromoacetyl and thiol oligonucleotide derivatives were covalently linked to sulfhydryl- and bromoacetyl-polyacrylamide supports with greater than 95% end-attachment efficiencies. These polyacrylamide-oligonucleotide supports were further derivatized with anionic residues to provide multi-functional supports which show low non-specific binding for non-complementary nucleic acids. While all the polyacrylamide-oligonucleotide supports capture complementary oligonucleotides with high affinity, the pore size was found to be a critical parameter in sandwich hybridization reactions. The superior hybridization characteristics of the Trisacryl support was ascribed to a combination of its macroporous nature, hydrophilicity and the terminal attachment of its capture oligonucleotides.     

Ethanol separation by selective adsorption of water

Preferential adsorption of water in the vapor phase by lignocellulosic residues for the production of anhydrous ethanol has been reported in this work. Rice straw, microcrystalline cellulose powder (MCCP) and bagasse were the lignocellulosic residues used as adsorbents. Starting with an ethanol concentration of 80–90% a final concentration above the azeotropic concentration was obtained. An energy analysis of the process was made and a possible explanation of phenomena suggested.     

Speed Breeding Opportunities and Challenges for Crop Improvement

Journal of Plant Growth Regulation - Crop improvement in light of the rapidly changing climate and the increasing human population continues to be one of the primary concerns for researchers across...     

Volatilization of mercury by immobilized mercury-resistant bacterial cells

Highly toxic mercury compounds may come into the environment through the use of mercury compounds as disinfectants for hospital and household purposes, Hg catalyst in industries, burning of coal and petroleum products, mercury-based pesticides and fungicides used in agriculture, and seed dressings. Toxic effects of mercury can be counteracted by microbial cells through the enzymes mercuric reductase and organomercurial lyase. Immobilized mercury-resistant bacterial cells of Azotobacter chroococcum could effectively volatilize mercury from mercury-containing buffer and detoxify mercury compounds. Moreover, the efficiency of mercury volatilization was much greater than with the native cells, as immobilized cells can be reused. Immobilized cells continuously volatilized mercury from mercury-containing buffer after four consecutive 24 h cycles. The storage stability of immobilized cells was much better than that of the native cells.     

Non-equilibrium thermodynamics of Lotka-Volterra ecosystems: Stability and evolution

A non-equilibrium thermodynamic theory of generalized Lotka-Volterra ecosystem has been presented. The main results consist of the derivation of a generalized expression of entropy-production for the evolutionary ecosystem and the study of its role in the analysis of ecological stability, succession and also in the formulation of some extremum principles characterising the evolution of the ecosystem.