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Automated cloning methods.

Description: Argonne has developed a series of automated protocols to generate bacterial expression clones by using a robotic system designed to be used in procedures associated with molecular biology. The system provides plate storage, temperature control from 4 to 37 C at various locations, and Biomek and Multimek pipetting stations. The automated system consists of a robot that transports sources from the active station on the automation system. Protocols for the automated generation of bacterial expression clones can be grouped into three categories (Figure 1). Fragment generation protocols are initiated on day one of the expression cloning procedure and encompass those protocols involved in generating purified coding region (PCR).
Date: August 22, 2001
Creator: Collart, F.
Partner: UNT Libraries Government Documents Department

Net assimilation and photosynthate allocation of Populus clones grown under short-rotation intensive culture: Physiological and genetic responses regulating yield

Description: The overall objective of this project was to determine the differential responses of poplar clones from sections Tacamahaca and Aigeiros of the genus Populus to varying levels of applied water and nitrogen. Above- and below-ground phenology and morphology, photosynthate allocation, and physiological processes were examined. By manipulating the availability of soil resources, we have been able to separate inherent clonal differences from plastic responses, and to determine genotype-environment interactions. We also have been able to make some contrasts between trees grown from hardwood cuttings and coppice sprouts. Our overall hypothesis was that carbon allocation during growth is greatly influenced by interactions among moisture, nitrogen, and genotype, and that these interactions greatly influence yield in short-rotation plantations. As is true of any project, some of our original expectations were not realized, whereas other initially unforeseen results were obtained. The reduced funding from the Biofuels Feedstock Development Program (BFDP) during the last few years of the project slowed us down to some extent, so progress was not been as rapid as we might have hoped. The major problem associated with this funding shortfall was the inability to employ skilled and unskilled student labor. Nonetheless, we were able to accomplish most of our original goals. All of the principal investigators on this project feel that we have made progress in advancing the scientific underpinning of short-rotation woody biomass production.
Date: August 1, 1996
Creator: Dickmann, D.I.; Pregitzer, K.S. & Nguyen, P.V.
Partner: UNT Libraries Government Documents Department

Viruses of eukaryotic green algae; Progress report, June 20, 1990--July 1, 1991

Description: Many large polyhedral, dsDNA containing (ca. 330 kb), plaque forming viruses which infect a unicellular, eukaryotic, chlorella-like green alga have been isolated and characterized. The plaque assay, the ability to synchronously infect the host, the short life cycle, and the ability of the viruses to undergo homologous recombination make them excellent model systems for studying many plant cell functions in the manner that bacterial and animal viruses have been used to study bacterial and animal cell functions. These viruses have several unique features including: (1) coding for DNA methyltransferase and site-specific (restriction) endonucleases and (2) unlike other viruses, these viruses appear to code for the enzymes involved in the glycosylation of their glycoproteins.
Date: December 31, 1991
Creator: Van Etten, J.L.
Partner: UNT Libraries Government Documents Department

Thermostabilization of desulfurization enzymes from Rhodococcos sp. IGTS8. Final technical report

Description: The objective of this project was to develop thermophilic cultures capable of expressing the desulfurization (dsz) operon of Rhodococcus sp. IGTS8. The approaches taken in this project included the development of plasmid and integrative expression vectors that function well in Thermus thermophilus, the cloning of Rhodococcus dsz genes in Thermus expression vectors, and the isolation of bacterial cultures that express the dsz operon at thermophilic temperatures. This project has resulted in the development of plasmid and integrative expression vectors for use in T. thermophilus. The dsz genes have been expressed at moderately thermophilic temperatures (52 C) in Mycobacterium phlei and at temperatures as high as 72 C in T. thermophilus. The tools and methods developed in this project will be generally useful for the expression of heterologous genes in Thermus. Key developments in the project have been the isolation of a Mycobacterium phlei culture capable of expressing the desulfurization operon at 52 C, development of plasmid and integrative expression vectors for Thermus thermophilus, and the development of a host-vector system based on the malate dehydrogenase gene that allows plasmids to be stably maintained in T. thermophilus and provides a convenient reporter gene for the accurate quantification of gene expression. Publications have been prepared regarding each of these topics; these preprints are included.
Date: December 15, 2000
Creator: II, John J. Kilbane
Partner: UNT Libraries Government Documents Department

Human Cloning

Description: This report discusses human cloning science and federal policy regarding human embryo research. It provides background on the topic, federal policies, state laws, Congressional actions, and ethical issues.
Date: July 20, 2006
Creator: Johnson, Judith A. & Williams, Erin D.
Partner: UNT Libraries Government Documents Department

Library Resources for Bac End Sequencing. Final Technical Report

Description: Studies directed towards the specific aims outlined for this research award are summarized. The RPCI II Human Bac Library has been expanded by the addition of 6.9-fold genomic coverage. This segment has been generated from a MBOI partial digest of the same anonymous donor DNA used for the rest of the library. A new cloning vector, pTARBAC1, has been constructed and used in the construction of RPCI-II segment 5. This new cloning vector provides a new strategy in identifying targeted genomic regions and will greatly facilitate a large-scale analysis for positional cloning. A new maleCS7BC/6J mouse BAC library has been constructed. RPCI-23 contain 576 plates (approx 210,000 clones) and represents approximately 11-fold coverage of the mouse genome.
Date: October 1, 2000
Creator: Jong, Pieter J. de
Partner: UNT Libraries Government Documents Department

Molecular and Genetic Analysis of Hormone-Regulated Differential Cell Elongation in Arabidopsis

Description: We have utilized the response of Arabidopsis seedlings to the plant hormone ethylene to identify new genes involved in the regulation of ethylene biosynthesis, perception, signal transduction and differential cell growth. In building a genetic framework for the action of these genes, we have developed a molecular model that has facilitated our understanding of the molecular requirements of ethylene for cell elongation processes. The ethylene response pathway in Arabidopsis appears to be primarily linear and is defined by the genes: ETR1, ETR2, ERS1, ERS2, EIN4, CTR1, EIN2, EIN3, EIN5, EIN6, and EIN. Downstream branches identified by the HLS1, EIR1, and AUX1 genes involve interactions with other hormonal (auxin) signals in the process of differential cell elongation in the hypocotyl hook. Cloning and characterization of HLS1 (and three HLL genes) and ETO1 (and ETOL genes) in my laboratory has been supported under this award. HLS1 is required for differential elongation of cells in the hypocotyl and may act in the establishment of hormone gradients. Also during the previous period, we have identified and characterized a gene that genetically acts upstream of the ethylene receptors. ETO1 encodes negative regulators of ethylene biosynthesis.
Date: September 15, 2005
Creator: Ecker, Joseph R.
Partner: UNT Libraries Government Documents Department

Technology development for gene discovery and full-length sequencing

Description: In previous years, with support from the U.S. Department of Energy, we developed methods for construction of normalized and subtracted cDNA libraries, and constructed hundreds of high-quality libraries for production of Expressed Sequence Tags (ESTs). Our clones were made widely available to the scientific community through the IMAGE Consortium, and millions of ESTs were produced from our libraries either by collaborators or by our own sequencing laboratory at the University of Iowa. During this grant period, we focused on (1) the development of a method for preferential cloning of tissue-specific and/or rare transcripts, (2) its utilization to expedite EST-based gene discovery for the NIH Mouse Brain Molecular Anatomy Project, (3) further development and optimization of a method for construction of full-length-enriched cDNA libraries, and (4) modification of a plasmid vector to maximize efficiency of full-length cDNA sequencing by the transposon-mediated approach. It is noteworthy that the technology developed for preferential cloning of rare mRNAs enabled identification of over 2,000 mouse transcripts differentially expressed in the hippocampus. In addition, the method that we optimized for construction of full-length-enriched cDNA libraries was successfully utilized for the production of approximately fifty libraries from the developing mouse nervous system, from which over 2,500 full-ORF-containing cDNAs have been identified and accurately sequenced in their entirety either by our group or by the NIH-Mammalian Gene Collection Program Sequencing Team.
Date: July 19, 2004
Creator: Soares, Marcelo Bento
Partner: UNT Libraries Government Documents Department

Characterization of Arabidopsis Genes Involved in Gene Silencing. Final Progress Report

Description: Enhancer of gene silencing 1 (egs1) is an Arabidopsis mutant that enhances post-transcriptional gene silencing of the rolB gene introduced by genetic engineering (transgene). The goal of our proposal was cloning EGS1 based on its map position. Although we screened more than 2000 chromosomes for recombination, we were unable to get closer than 2 cM to the gene. We experienced an unexpected tendency of the post-transcriptionally silenced transgene to switch to a more stable silenced state. This made it impossible to select egs1 homozygotes for map based cloning. This forced us to reconsider our cloning strategy. One possibility would have been to use a different transgene as the target of gene silencing. We tested two other transgenes. Both encoded proteins unrelated to the first but they were all expressed from the same type of promoter and they all had a similar tendency to become post-transcriptionally silenced. After screening over 80 F2 segregants from each cross between our egs1 mutant and Arabidopsis of the same ecotype homozygous for the new transgene, we were disappointed to find that the egs1 mutation did not enhance post-transcription silencing of the two new genes. In 80 plants we expected to have between 4 and 6 plants that were homozygous for the transgene and for the mutant egs1 allele. If egs1 mutations could enhance gene silencing of the new transgene, these plants would not express it. However all the double homozygotes still expressed the transgene. Therefore, we could not change the target transgene for mapping. This was the state of the cloning at the time for renewal of the grant in 1999. Because the selection of new meaningful recombinant plants had become extremely inefficient using the original rolB transgene, we abandoned the attempt at map based cloning and did not apply for further funding.
Date: February 5, 1999
Creator: Grant, S. R.
Partner: UNT Libraries Government Documents Department

A nanostructure-initiator mass spectrometry-based enzyme activity assay

Description: We describe a Nanostructure-Initiator Mass Spectrometry (NIMS) enzymatic (Nimzyme) assay in which enzyme substrates are immobilized on the mass spectrometry surface by using fluorous-phase interactions. This 'soft' immobilization allows efficient desorption/ionization while also enabling the use of surface-washing steps to reduce signal suppression from complex biological samples, which results from the preferential retention of the tagged products and reactants. The Nimzyme assay is sensitive to subpicogram levels of enzyme, detects both addition and cleavage reactions (sialyltransferase and galactosidase), is applicable over a wide range of pHs and temperatures, and can measure activity directly from crude cell lysates. The ability of the Nimzyme assay to analyze complex mixtures is illustrated by identifying and directly characterizing {beta}-1,4-galactosidase activity from a thermophilic microbial community lysate. The optimal enzyme temperature and pH were found to be 65 C and 5.5, respectively, and the activity was inhibited by both phenylethyl-{beta}-d-thiogalactopyranoside and deoxygalactonojirimycin. Metagenomic analysis of the community suggests that the activity is from an uncultured, unsequenced {gamma}-proteobacterium. In general, this assay provides an efficient method for detection and characterization of enzymatic activities in complex biological mixtures prior to sequencing or cloning efforts. More generally, this approach may have important applications for screening both enzymatic and inhibitor libraries, constructing and screening glycan microarrays, and complementing fluorous-phase organic synthesis. The interest in leveraging mass spectrometry for studying enzyme activities in complex biological samples derives from its high sensitivity and specificity; however, signal suppression and significant sample preparation requirements limit its overall utility (1). Here we describe a Nanostructure-Initiator Mass Spectrometry (NIMS) enzymatic (Nimzyme) assay, which uses the fluorous liquid-coated surface of NIMS (2) to noncovalently attach enzyme substrates by means of fluorous tags. Enzymes play essential roles in a wide range of cellular processes and account for >20% of all drug targets (3). In addition, enzymes have ...
Date: March 10, 2008
Creator: Siuzdak, Gary; Northen, Trent R.; Lee, Jinq-Chyi; Hoang, Linh; Raymond, Jason; Hwang, Der-Ren et al.
Partner: UNT Libraries Government Documents Department

Use of bromodeoxyuridine immunocapture to identify psychrotolerant phenanthrene-degrading bacteria in phenanthrene-enriched polluted Baltic Sea sediments

Description: The aim of this study was to enrich and identify psychrotolerant phenanthrenedegrading bacteria from polluted Baltic Sea sediments. Polyaromatic hydrocarbon (PAH)-contaminated sediments were spiked with phenanthrene and incubated for 2 months in the presence of bromodeoxyuridine that is incorporated into the DNA of replicating cells. The bromodeoxyuridine-incorporated DNA was extracted by immunocapture and analyzed by terminal-restriction fragment length polymorphism and 16S rRNA gene cloning and sequencing to identify bacterial populations that were growing. In addition, degradation genes were quantified in the bromodeoxyuridine-incorporated DNA by real-time PCR. Phenanthrene concentrations decreased after 2 months of incubation in the phenanthrene-enriched sediments and this reduction correlated to increases in copy numbers of xylE and phnAc dioxygenase genes. Representatives of Exiguobacterium, Schewanella,Methylomonas, Pseudomonas, Bacteroides and an uncultured Deltaproteobacterium and a Gammaproteobacterium dominated the growing community in the phenanthrene spiked sediments. Isolates that were closely related to three of these bacteria (two pseudomonads and an Exiguobacterium sp.) could reduce phenanthrene concentrations in pure cultures and they all harbored phnAc dioxygenase genes. These results confirm that this combination of culture-based and molecular approaches was useful for identification of actively growing bacterial species with a high potential for phenanthrene degradation.
Date: May 1, 2008
Creator: Edlund, A. & Jansson, J.
Partner: UNT Libraries Government Documents Department

Structural Genomics of Minimal Organisms: Pipeline and Results

Description: The initial objective of the Berkeley Structural Genomics Center was to obtain a near complete three-dimensional (3D) structural information of all soluble proteins of two minimal organisms, closely related pathogens Mycoplasma genitalium and M. pneumoniae. The former has fewer than 500 genes and the latter has fewer than 700 genes. A semiautomated structural genomics pipeline was set up from target selection, cloning, expression, purification, and ultimately structural determination. At the time of this writing, structural information of more than 93percent of all soluble proteins of M. genitalium is avail able. This chapter summarizes the approaches taken by the authors' center.
Date: September 14, 2007
Creator: Kim, Sung-Hou; Shin, Dong-Hae; Kim, Rosalind; Adams, Paul & Chandonia, John-Marc
Partner: UNT Libraries Government Documents Department

Statistical methods in physical mapping

Description: One of the great success stories of modern molecular genetics has been the ability of biologists to isolate and characterize the genes responsible for serious inherited diseases like fragile X syndrome, cystic fibrosis and myotonic muscular dystrophy. This dissertation concentrates on constructing high-resolution physical maps. It demonstrates how probabilistic modeling and statistical analysis can aid molecular geneticists in the tasks of planning, execution, and evaluation of physical maps of chromosomes and large chromosomal regions. The dissertation is divided into six chapters. Chapter 1 provides an introduction to the field of physical mapping, describing the role of physical mapping in gene isolation and ill past efforts at mapping chromosomal regions. The next two chapters review and extend known results on predicting progress in large mapping projects. Such predictions help project planners decide between various approaches and tactics for mapping large regions of the human genome. Chapter 2 shows how probability models have been used in the past to predict progress in mapping projects. Chapter 3 presents new results, based on stationary point process theory, for progress measures for mapping projects based on directed mapping strategies. Chapter 4 describes in detail the construction of all initial high-resolution physical map for human chromosome 19. This chapter introduces the probability and statistical models involved in map construction in the context of a large, ongoing physical mapping project. Chapter 5 concentrates on one such model, the trinomial model. This chapter contains new results on the large-sample behavior of this model, including distributional results, asymptotic moments, and detection error rates. In addition, it contains an optimality result concerning experimental procedures based on the trinomial model. The last chapter explores unsolved problems and describes future work.
Date: May 1, 1995
Creator: Nelson, D.O.
Partner: UNT Libraries Government Documents Department

Armored Enzyme Nanoparticles for Remediation of Subsurface Contaminants

Description: The remediation of subsurface contaminants is a critical problem for the Department of Energy, other government agencies, and our nation. Severe contamination of soil and groundwater exists at several DOE sites due to various methods of intentional and unintentional release. Given the difficulties involved in conventional removal or separation processes, it is vital to develop methods to transform contaminants and contaminated earth/water to reduce risks to human health and the environment. Transformation of the contaminants themselves may involve conversion to other immobile species that do not migrate into well water or surface waters, as is proposed for metals and radionuclides; or degradation to harmless molecules, as is desired for organic contaminants. Transformation of contaminated earth (as opposed to the contaminants themselves) may entail reductions in volume or release of bound contaminants for remediation. Research at Rensselaer focused on the development of haloalkane dehalogenase as a critical enzyme in the dehalogenation of contaminated materials (ultimately trichloroethylene and related pollutants). A combination of bioinformatic investigation and experimental work was performed. The bioinformatics was focused on identifying a range of dehalogenase enzymes that could be obtained from the known proteomes of major microorganisms. This work identified several candidate enzymes that could be obtained through relatively straightforward gene cloning and expression approaches. The experimental work focused on the isolation of haloalkane dehalogenase from a Xanthobacter species followed by incorporating the enzyme into silicates to form biocatalytic silicates. These are the precursors of SENs. At the conclusion of the study, dehalogenase was incorporated into SENs, although the loading was low. This work supported a single Ph.D. student (Ms. Philippa Reeder) for two years. The project ended prior to her being able to perform substantive bioinformatics efforts that would identify more promising dehalogenase enzymes. The SEN synthesis, however, was demonstrated to be partially successful with dehalogenases. ...
Date: February 19, 2007
Creator: Dordick, Jonathan S.; Grate, Jay & Kim, Jungbae
Partner: UNT Libraries Government Documents Department

Genes for Uranium Bioremediation in the Anaerobic Sulfate-Reducing Bacteria

Description: Objective A: Electron transfer components necessary for uranium reduction. Objective B: Possible FNR-analog in the sulfate-reducing bacteria. Attempts to isolate FNR or FIKJ analogs from Desuflovibrio through the design of degenerate primers for amplification of portions of the genes has not been successful. In contrast, several amplicons have been generated for the genes encoding the regulators of two-component signal sequences. Since several global regulators fall into this class, we are attempting to obtain sufficient sequence information to indicate what metabolic pathways are affected by the regulators. Cloning and sequencing of two such amplicons has revealed that bona fide two-component regulators are present in Desulfovibrio.
Date: June 1, 1999
Creator: Wall, Judy D.
Partner: UNT Libraries Government Documents Department

Enzymatic Upgrading of Heavy Crudes via Partial Oxidation or Conversion of PAHs

Description: The objective of this program was to investigate new enzyme-based technologies for upgrading of heavy oils. Enzymes were selected for screening from those capable of conversion of polyaromatic hydrocarbons (PAHs) reported in the literature. Oxidative reactions of PAHs using hydrogen peroxide as an oxidant with conversion to partially oxidized products were used. The enzymes (lignin peroxidase, cytochrome c) were tested in various organic solvents and found to loose activity in pure organic solvents. A thermodynamic analysis revealed lack of effective interaction between the substrate and enzyme as the cause for low activity. The protein cytochrome c was modified to work in organic media by chemical hydrophobic group attachment. Two different modifications were made: attachment of polyethylene glycol (PEG) and alkyl groups. Alkyl groups, being small could be attached at interior locations within the core of the enzyme and possibly near the active site. Increase in the threshold solvent concentration where maximum enzyme activity occurred indicated potential of this strategy for effective enzyme-substrate interaction. Further improvements in enzyme activity called for other diverse methods due to the unavailability of sufficient chemical modification sites. Genetic techniques were therefore explored for further improvements. These experiments focused on cloning of a gene for the fungal enzyme lignin peroxidase (lip) into yeast Pichia pastoris, which would allow easy manipulation of the gene. However, differences in the fungal and yeast cellular machinery impeded significant expression of the fungal enzyme. Several strategies were explored to allow higher-level expression of the enzyme, which was required for enzyme improvement. The strategies used in this investigation are described in the report. Industrial in-kind support was available throughout the project period. review of the research results was carried out on a regular basis (bimonthly reports and annual meetings) followed by suggestions for improvement in ongoing work and direction for future work. ...
Date: July 1, 2002
Creator: Borole, A.P.; Davison, B.H. & Kuritz, T.
Partner: UNT Libraries Government Documents Department

Molecular characterization of flow-sorted mammalian centromeres

Description: This is the final report of a three-year, Laboratory Directed Research and Development (LDRD) project at the Los Alamos National Laboratory (LANL). The project involved experiments directed towards developing a molecular characterization of the centromere region of mammalian chromosomes. Attempts to purify this essential chromosomal locus by conventional methods have thus far been unsuccessful. However, preliminary data obtained in collaboration with the National Flow Cytometry Resource (NFCR) showed that it is possible to purify a chromosome fragment that is present in certain cultured mouse cell lines and has all the properties expected of an intact centromere region. To begin sorting this minichromosome for the identification of proteins preferentially associated with centromere regions, standard buffers utilized in chromosome sorting were evaluated for potential effects on maintenance of chromosomal proteins during sorting. The data indicate that the presence of several buffer constituents results in the extraction of all but a few chromosomal proteins. The subsequent use of a magnesium sulfate buffer resulted in the sorting of mouse chromosomes that do not suffer a significant loss of proteins. Several DNA stains were also evaluated for causing protein dissociation, but no significant losses were observed. Although flow-sorted chromosomes have been used extensively for DNA analysis and cloning, this is a pioneering effort by the NFCR, and its collaborators, to exploit chromosome sorting capabilities for the analysis of chromosomal proteins.
Date: December 31, 1998
Creator: Hamkalo, B.A.; Henschen, A. & Parseghian, M.H.
Partner: UNT Libraries Government Documents Department

Molecular Profiling of Microbial Communities from Contaminated Sources: Use of Subtractive Cloning Methods and rDNA Spacer Sequences

Description: The major objective of the research was to provide appropriate sequences and to assemble a DNA arrays of oligonucleotides to be used for rapid profiling of microbial populations, from polluted areas and from areas of other interest. The sequences to be assigned to the DNA array are chosen from cloned genomic DNA from groundwater at DOE sites containing organic solvents. The sites, Hanford Nuclear Plant and Lawrence Livermore Site 300 (LLNL), have well characterized pollutant histories, which have been provided by our collaborators.
Date: June 1, 2000
Creator: Robb, Frank T.
Partner: UNT Libraries Government Documents Department

Progress Report: DE-FG03-97ER20274, ''Microbial Production of Isoprene''

Description: We have discovered that microorganisms produce and emit the hydrocarbon isoprene (2-methyl-1,3-butadiene), and have suggested that if isoprene-producing enzymes and their genes can be harnessed, useful hydrocarbon-producing systems might be constructed. The main goal of the proposed work is to establish the biochemical mechanism and regulation of isoprene formation in the bacterial system, Bacillus subtilis. Specific objectives of the proposed work are the following: (A) to characterize the physiological regulation of isoprene formation in B. subtilis; (B) to characterize mutations in B. subtilis 168 that suppress isoprene formation, clone these genes, and determine how isoprene and isoprenoid carbon flow are regulated; and (C) to test ''overflow'' and ''signaling'' models for Bacillus isoprene formation. We are also pursuing the isolation and cloning of B. subtilis isoprene synthase, which we believe may be a regulatory enzyme.
Date: March 13, 2002
Creator: Fall, Ray
Partner: UNT Libraries Government Documents Department

Hemicellulases from the ethanologenic thermophile Thermoanaerobacter ethanolicus and related anaerobic thermophiles. Final report, September 1992--June 1996

Description: The SHORT TERM GOALS of this application were to characterize hemicellulases from anaerobic thermophiles on the biochemical and molecular level to extend the presently limited knowledge of hemicellulases in anaerobic thermophilic bacteria. This objective includes the following TASKS: (1) Traditional purification and biochemical/biophysical characterization of xylanases from the newly isolated, slightly alkalitolerant strain NDF190, and the slightly acid-tolerant strain YS485, both with high xylanolytic activities, and of the 4-0-methyl glucuronidase and arabinosidase from strain NDF190 and the acetyl (xylan) esterase from T. ethanolicus. This also includes determining the N-terminal sequences and obtaining gene probes. (2) Elucidation of the regulation of hemicellulolytic enzymes in anaerobic thermophiles. (3) To clone into E. coli and identify the multiplicity of the enzymes involved in hemicellulose degradation by T. ethanolicus and other suitable organisms. (4) To purify and characterize the recombinant enzymes with the goal of identifying the best enzymes for cloning into the ethanologenic T. ethanolicus to obtain an optimized hemicellulose utilization by this bacterium (one of our long term goals).
Date: May 1, 1998
Creator: Wiegel, J.
Partner: UNT Libraries Government Documents Department

Genetics and molecular biology of methanogen genes. Final report

Description: Adenylate kinase has been isolated from four related methanogenic members of the Archaea. For each the optimum temperature for enzyme activity was similar to the temperature for optimal microbial growth and was approximately 30 C for Methanococcus voltage, 70 C for Methanococcus thermolithotrophicus, 80 C for Methanococcus igneus and 80--90 C for Methanococcus jannaschii. The enzymes were sensitive to the adenylate kinase inhibitor, Ap{sub 5}A [P{sup 1}, P{sup 5}-di(adenosine-5{prime}) pentaphosphate], a property that was exploited to purify the enzymes by CIBACRON Blue affinity chromatography. The enzymes had an estimated molecular weight (approximately 23--25 kDa) in the range common for adenylate kinases. Each of the enzymes had a region of amino acid sequence close to its N-terminus that was similar to the canonical P-loop sequence reported for all adenylate kinases. However, the methanogen sequences lacked a lysine residue that has previously been found to be invariant in adenylate kinases including an enzyme isolated from the Archeon, Sulfolobus acidocaldarius. If verified as a nucleotide binding domain, the methanogen sequence would represent a novel nucleotide binding motif. There was no correlation between amino acid abundance and the optimal temperature for enzyme activity.
Date: October 7, 1997
Creator: Konisky, J.
Partner: UNT Libraries Government Documents Department

Large scale solubilization of coal and bioconversion to utilizable energy. Quarterly report, October 1--December 31, 1996

Description: In order to develop a system for a large scale coal solubilization and its bioconversion to utilizable fuel, the author plans to clone the genes encoding Neurospora protein that facilitate depolymerization of coal. He also plans to use desulfurizing bacteria to remove the sulfur in situ and use other microorganisms to convert biosolubilized coal into utilizable energy following an approach utilizing several microorganisms. In addition the product of coal solubilized by fungus will be characterized to determine their chemical nature and the mechanism of reaction catalyzed by fungal product during in vivo and in vitro solubilization by the fungus or purified fungal protein.
Date: December 22, 1996
Creator: Mishra, N.C.
Partner: UNT Libraries Government Documents Department

Increasing the productivity of biomass plantations of Populus species and hybrids in the Pacific Northwest. Final report, September 14, 1981--December 31, 1996

Description: This final report represents the culmination of eight years of biological research devoted to increasing the productivity of short rotation plantations of Populus trichocarpa and Populus hybrids in the Pacific Northwest. Studies described herein provide an understanding of tree growth, stand development and biomass yield at various spacings, and how patterns thereof differ by Populus clone in monoclonal and polyclonal plantings. Also included is some information about factors related to wind damage in Populus plantings, use of leaf size as a predictor of growth potential, and approaches for estimating tree and stand biomass and biomass growth. The work was accomplished in three research plantations, all established cooperatively with the Washington State Department of Natural Resources (DNR) and located at the DNR Tree Improvement Center near Olympia. The first plantation was established in Spring 1986 to evaluate the highly touted {open_quotes}woodgrass{close_quotes} concept and compare it with more conventional short-rotation management regimes, using two Populus hybrid clones planted at five spacings. Besides providing scientific data to resolve the politicized {open_quotes}wood-grass{close_quotes} dispute, this plantation has furnished excellent data on stand dynamics and woody biomass yield. A second plantation was established at the same time; groups of trees therein received two levels of irrigation and different amounts of four fertilizer amendments, resulting in microsites with diverse moisture and nutrient conditions.
Date: August 1, 1997
Creator: DeBell, D.S.; Harrington, C.A. & Clendenen, G.W.
Partner: UNT Libraries Government Documents Department

Hydrogen from renewable resources monthly progress report

Description: During February, we achieved two significant results in our hydrogen storage activates. Reversible hydrogen uptake and release was measured at room temperature, near ambient pressure on the (IrClH{sub 2}(H{sub 2})Pr{sup i}{sub 3}) complex. Dr. Jensen also observed that certain polyhydzide complexes catalyze the low temperature, reversible dehydrogenation of cycloalkanes to aromatic hydrocarbons at temperatures as low as 130{degrees}C. This discovery may represent a breakthrough in chemical storage of hydrogen as all other cycloalkane dehydrogenation systems require temperatures in excess of 300{degrees}C.
Date: February 1, 1995
Creator: Rocheleau, R.E.
Partner: UNT Libraries Government Documents Department