Browsing by Subject "Deoxyribonucleic acid (DNA)"
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Item Analysis of DNA-Binding Nonhistone Chromosomal Proteins(Texas Tech University, 1978-12) Lee, Shuey-ChyongNot Available.Item Applications of affinity chromatography to isolation of sequence specific nuclear proteins(Texas Tech University, 1981-05) Sherrod, Peter D.Even before Watson and Crick proposed their brilliant model for the structure of DNA, Jacob and Monod were busy piecing together the information that would lead to the first fully defined mechanism for genetic regulation in the bacteria, Escherichia coli. Since then, other mechanisms have become fell defined in prokaryotes, however eukaryotic mechanisms of control have been difficult to establish. This is primarily due to the fact that the genome of euiiaryotic cells is much more complex than in bacteria. Just the immensity of size defies the precise and controlled study that has yielded so much information in prokaryotic systems. Nevertheless, it is not inconceivable that similar mechanisms of regulation exist in the eukaryotic genome.Item Characterization of an ORF in a novel orientation in Escherichia coli(Texas Tech University, 2003-12) Lackey, Melinda K.Evidence has been presented in the literature that demonstrates the expendable nature of the C-terminal region for the proper functioning of TolC in the bacterial cell. Recent sequencing of various microbial genomes demonstrated the presence of 3' overlapping, converging open reading frames involving tolC and its homologues found in other species and genera of bacteria. In this study, we examined the Escherichia coli tolC gene and the 3' overlapping, converging ORF found at the same genetic locus, denoted cloT for tolC in reverse. The expression of c/or was examined using reverse transcriptase PCR to identify the presence of mRNA molecules within the bacterial cell. Amplification of a 120 base pair fragment from total RNA isolated from wild-type E. coli suggested that cloT was transcribed, at least at low levels, in the organism. To corroborate these data, the upstream region of the structural gene was analyzed for the ability to drive transcription of a reporter gene, beta-galactosidase. Activity analysis of this reporter fusion demonstrated the ability of the upstream region of the cloT gene to drive transcription in E. coli. Similarly, we have demonstrated an interaction between the upstream region of cloT and soluble proteins isolated from the cytoplasm of wild-type E. coli. Further experiments were attempted to raise antibodies against a portion of the putative CloT protein using both GST fusion technology and a synthetic peptide. Neither has been fully characterized at time of publication. In addition, attempts to isolate mutations in the unique region of the cloTORF in the chromosome of E. coli were also unsuccessful. Examination of the information known about TolC and its homologues in different genera of bacteria revealed interesting information concerning the C-termini of these proteins. Alignment of the amino acid sequences demonstrated lower identity shared in the C-terminal region of these proteins. In addition, examination of the nucleotide sequences of these homologues indicated the presence of a similar open reading frame sharing a convergent, overlapping relationship with its respective tolC homologue. When combined with published data concerning the expendable nature of the last fifty amino acids of TolC, these data suggest that the presence of a conserved open reading frame in this region may play an important role in the regulation or function of the tolC gene or gene product(s).Item Development of species- and genome-specific genetic markers by representational difference analysis: application in systematic and evolutionary research(Texas Tech University, 1999-08) Nekrutenko, AntonIn the study of biodiversity, it is important to have a reliable system for identification of various genetically distinct units (species, subspecies, etc). One of the most efficient tools available today is the polymerase chain reaction (PCR) with diagnostic primers, that yield a detectable product for one taxon but not for other taxa. Critical to this method is the identification of diagnostic DNA fragments from which primers can be designed. Representational difference analysis (RDA) can reliably isolate DNA fragments that are unique to a specific taxon. In this report, we demonstrate the utility of the technique by development of binary markers that distinguish between two cryptic species of voles (genus Microtus).Item DNA transformation and mutagenesis studies on dithane with Bacillus subtilis repair-deficient strains(Texas Tech University, 1980-08) Lee, Ming-chouNot availableItem DNA-binding proteins in Escherichia coli(Texas Tech University, 1977-05) Eaton, Leslie C.Not availableItem Functional characterization of transcriptional inhibitory domains in the C/EBP-epsilon basic region/leucine zipper transcription factor(Texas Tech University, 2003-08) Li, YaminC/EBPE is a member of the CCAAT/enhancer binding protein (C/EBP) family of transcription factors and is primarily expressed in neutrophils. Multiple functional domains were previously identified in the C/EBPe polypeptide including a regulatory domain (RD or RDI) that negatively regulates the activity of C/EBPe. Within the RD a five amino acid motif (the regulatory domain motif (RDM)) is conserved in three other C/EBP family members and is critical for the inhibitory function of the RD. Interestingly the RDM is similar to the recognition sequence for the small ubiquitin-like modifier protein (SUMO) and attachment of SUMO to the RDM can activate C/EBPe. The goal of this study was to explore the molecular mechanisms by which the RD controls C/EBPe activity. These experiments may provide important information about general transcriptional regulatory mechanisms as RD-like elements, and SUMO attachment sites, exist in inhibitory domains of a large number of transcriptional regulators. Two models were designed to explain the mechanism of the function of the RD. One model proposes that intra-molecular interactions exist between the RD and its linked activation domain (AD), thereby blocking access of the AD to components of the transcriptional machinery. In this model, the attachment of SUMO to the RDM releases the AD and activates C/EBPe. Physical interactions between the RD and AD were not detected in either a mammalian two-hybrid assay or a direct interaction assay. The validity of this first model was also assessed by examining the activation domain specificity of the RD. The C/EBPe RD was capable of inhibiting linked ADs from three different classes, suggesting that the RD is unlikely to function according to model one. The second model assumes the RD functions by recruiting as yet unknown inhibitory RDM binding factor(s) (RDM-BF). In this model, SUMO attachment would displace the RDM-BFs thereby releasing the inhibitory effect of the RD. The first candidate RDM-BF tested was the family of proteins with histone deacetylase (HDAC) activity as these proteins are classically associated with co-repressor activity. Although the activity of C/EBPe was increased in the presence of the general HDAC inhibitor Trichostatin A, this effect was not dependent on the integrity of the RDM. These results indicate that C/EBPe may recruit HDACs through a domain separate from the RDM. In conclusion, the AD and RD of C/EBPe appear not to directly interact, however, additional candidate RDM-BFs must be identified and examined to determine whether the RDM functions by recruiting accessory, inhibitory proteins.Item Further purification of DNA-binding nonhistone chromosomal proteins(Texas Tech University, 1977-08) Tsai, SueshunNot availableItem Isolation and characterization of DNA sequences bound by a class of nonhistone proteins(Texas Tech University, 1979-08) Jagodzinski, Linda L.All somatic cells of the same organism contain the same complement of genes. During cellular differentiation transcriptional specialization occurs. This process allows the selected expression of genetic information in specialized cells; e.g., only red blood cell precursors synthesize hemoglobin, only hepatocytes synthesize phenylalanine hydroxylase and serum albumin (159), and only estrogen induced oviduct cells synthesize ovalbumin. During differentiation certain genes function only at specific times and in particular tissues. Hence, portions of the eukaryotic genome must be prevented from expressing, in some manner, their genetic information. Evidence indicates that the chromosomal proteins participate in the regulation of gene activity. How this is accomplished and which components are involved are questions which are now being investigated.Item Parallelization Methods for the Distribution of High-Throughput Bioinformatics Algorithms(2011-05) Rees, Eric; Youn, Eunseog; Dowd, Scot E.; San Francisco, MichaelThe development of high-throughput bioinformatics technologies has caused a massive influx of biological data over the course of the past decade. During this same span of time, computational hardware has also been rapidly increasing in speed while decreasing in price, multi-core processors have become standard in home and office environments, and distributed and cloud based computing has become affordable and readily available to researchers with implementations such as Amazon’s S3, Microsoft’s Azure, Google’s App Engine, and the 3Tera Cloud. Bioinformatics software tools such as BLAST, a tool for finding local alignments between a set of unknown genetic sequences versus a set of known genetic sequences, have simple interfaces and few installation requirements often so biologists can use them easily in the laboratory without needing an in-depth knowledge of how computer systems work. This, however, is rarely the case for distributed implementations of bioinformatics tools which often require the user to first set up and configure the underlying program that will handle the distribution, such as the Message Passing Interface (MPI). Once the underlying distribution algorithm is chosen, many of the software tools require the user to then configure the program to work with their chosen method and, in some cases, write the necessary source code to link the program with the underlying service. These are difficult steps for most computer scientists and are near impossible for the average biologist. By constructing a modularized set of methods that can connect to, broadcast to, and read from a multicast created by the methods, future bioinformatics software developers will be able to construct the underlying message passing system without requiring the end-user, often a biologist, to set up and configure one of their own. Using these multicast methods will allow any program the ability to seek out and track any nodes on the network that will be used in the distributed system. This communication method allows the program to easily scale up and down depending on available nodes without direct user intervention to alter the size of the system. This system is then tested by creating a program that connects NCBI’s Basic Local Alignment Search Tool to the multicast system to allow the BLAST algorithm to then be distributed across multiple nodes. This new system will demonstrate how future programs could then connect stand alone tools, such as BLAST, to the multicast system to create programs that will execute on a distributed system and automatically scale depending on the network size without altering the tools source code.Item Persistence of Adh gene in the environment, in Drosophila pupae, and in the mite Proctolaelaps regalis(Texas Tech University, 1997-05) Chasteen, Leslie A.There is substantial evidence supporting the horizontal gene transfer of DNA (P mobile elements) from Drosophila willistoni to Drosophila melanogaster. A potential vector of this transfer is the mite F^octolaelaps regalis. The mite is hypothesized to acquire DNA while feeding on flies and to carry and transmit that DNA to subsequent prey. This research concerns the persistence and longevity of DNA in the respective environmental components. Separate experiments were run testing: (1) the persistence of DNA in the environment using agar blocks as the substratum, (2) the persistence of DNA in Drosophila pupae after microinjection of the DNA, and (3) the persistence of DNA in the mite, Proctolaelaps regalis after feeding. A surrogate for the P element was used in the experiments in the form of a recombinant plasmid with a mammaUan alcohol dehydrogenase insert (Adh-V). The Adh-V signal was detected in the environment up to and including 14 days, in developing pupae up to the time of emergence and in emerged adults, and in the mite up to and including 60 hours. The data generated from these experiments provides estimates of the time frame in which a mite can potentially acquire, retain, and be competent to transfer genetic material among eukaryotes upon which it feeds.Item Photoinduced interaction of 8-methoxypsoralen (a skin sensitizer) with DNA and its effect on DNA replication(Texas Tech University, 1977-08) Tsai, Chee-HwayNot availableItem Screening pesticides with a colorimetric phage induction essay(Texas Tech University, 1981-05) Hume, Sharlene HelenNot availableItem Temperature dependent release of BB' subunits of DNA-dependent RNA polymerase from folded nucleoids of Escherichia coli dnaA [superscript] ts mutants and location of the 1c gene of bacteriophage PA-2(Texas Tech University, 1980-05) Eaton, Leslie C.Not availableItem The feeding and reproductive behavior of Proctolaelaps regalis (Gamasina: Ascidae)(Texas Tech University, 1999-05) London, Darin M.Proctolaelaps regalis DeLeon (Acari: Ascidae) is a species of free-living, omnivorous mites recently implicated as a potential vector in a horizontal gene transfer event between Drosophila willistoni and D. melanogaster. In this research, I investigated behaviors of the mite impacting its potential to acquire, maintain within the mite population, and transfer genes between flies. The potential of the mite to acquire and transfer genes is related to its propensity to prey upon the fly, but also to the population size and density of mites and flies within their habitat. Thus, aggregation and social behaviors exhibited by mites should influence their potential to acquire and transfer genes, and to maintain genes within the population. Mites were observed clustering in groups within the colony and depositing eggs in clusters. Mites were also observed to prefer fly growth medium previously conditioned to conspecific mites in culture over fresh medium, suggesting the presence of an aggregation pheromone. To investigate the propensity of the mites to prey upon D. melanogaster, and the search behavior response of mites to conspecific eggs and conspecific-conditioned growth medium, I used a paired-sample design in which individual mites were sequentially exposed to a control environment and seven potential resources. Each mite's behavior was recorded to determine its response. Mites were assessed for general attraction to the treatment in relation to their behavior in the control environment. Mites were also assessed for shifts in their search path pattern in response to the treatments. Mites were observed in small colonies for behaviors not addressed in the research design. The behavior of the mite was not influenced by the life-history stages of D. melanogaster. Mites were attracted to conspecific eggs, and strongly attracted to the conditioned growth medium. Mites exhibited potential social interactions within the colony, including intimate contacts between individual mites. They also exhibited aggressive behaviors and egg cannibalism. The lack of attraction to flies could decrease the mite's potential to acquire and transmit the gene. However, the mites may prey upon flies opportunistically, or switch to predation under periods of habitat decline. This would limit predation to certain times, but would not necessarily reduce the mite's potential as a vector.Item The immune response of CF-1 mice to single stranded DNA.(Texas Tech University, 1974-12) Reeves, James PattonNot availableItem The interaction of 4-nitroquinoline 1-oxide with Bacillus subtilis deoxyribonucleic acid(Texas Tech University, 1970-12) Laumbach, Andrew DeitliefNot availableItem The isolation and characterization of partial cDNAs associated with In Vitro tracheary element formation(Texas Tech University, 1995-08) Koonce, Linda TDifferential Display provides a sensitive method for quickly isolating and cloning cDNAs with corresponding mRNA transcripts that are differentially expressed between two different tissues or cell t5rpes. This technique provided a means by which three cDNAs with corresponding transcripts that are strongly expressed in suspension cultures of Zinnia mesophyll cells differentiating into tracheary elements were isolated. One transcript (63.30) has similarity to the non-histone protein 2 high mobility group-like protein in yeast, and to ribosomal proteins from yeast and the PRL2 protein from A. thaliana. The 63.30 transcript was found to be expressed in newly forming tracheary elements and phloem fibers and is expressed in response to auxin and/or cytokinin. A second transcript (54.6) was also determined to be expressed in newly forming tracheary elements and phloem fibers and is expressed in response to auxin and/or cytokinin. The deduced protein sequence of the 54.6 transcript shares some similarity to membrane spanning domains and the voltage sensor domain of the alpha subunit of the dihydropyridine- sensitive voltage-dependent calcium channel isolated from rat aorta. A third cDNA was isolated and expression of the transcript was found in in vitro differentiating tracheary elements, but no expression was detected in any whole plant tissue. It is possible that this transcript is a very rare species or is a result of the stress of culturing. No similarities to this transcript were detected by searches of the Blast/NCBI databases.Item The organization and abundance of repetitive DNA sequences among great apes and humans(Texas Tech University, 1995-08) Powell, Madison S.Most studies of the human genome are focused upon coding DNA sequences which provide the structural genes and enzymatic machinery necessary for life. However, repetitive DNA sequences constitute a substantial portion of eukaryotic genomes. Formerly referred to as "junk" DNA, their importance is just now being realized as many of these repetitive sequences have been implicated in human disease, chromosomal structure, and gene regulation. Our view of human genome organization and human evolution is incomplete, without an understanding of how repetitive DNA has evolved to its present abundance and distribution. To address this problem genomic libraries were constructed from DNA samples of a human (Homo sapiens) and representatives from each of the four extant great apes; gorilla (Gorilla gorilla) chimpanzee (Pan troglodvtes)bonobo Pan paniscus). and orangutan (Pongo pvgmaeus). The libraries were screened with radiolabeled, synthetic oligonucleotides for the presence of the major classes of repetitive DNAs found in humans. These repetitive classes included tandemly repeated microsatellites, a minisatellite. and satellite DNAs as well as dispersed repetitive sequences represented by a LINE-1 element, a SINE, and a human retrotransposon, THE1. Addressing genome organization with this technique has proven useful because it provided a minimal estimate of human repetitive element copy number similar to the estimates obtained in previous studies through examination of genomic and chromosome 16 specific DNA libraries. However, the abundance of some repetitive probes varied considerably among the ape libraries, indicating variation in repetitive element copy number can be substantial among closely related species. The technique also indicated non-random associations between selected repetitive sequences and supported a previously erected hypothesis that a-satellite DNA and Alu sequences are negatively associated.Item The protection of recombinationless Bacillus subtilis from four-nitroquinoline-one-oxide treatment by infection with bacteriophage PBS-1(Texas Tech University, 1970-12) Strong, James EvansNot available