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  Partner: UNT Libraries
 Department: Department of Biological Sciences
Stress Response by Alternative Σ-factor, Rpoh, and Analysis of Posttranslational Modification of the Heat Shock Protein, Dnak, in Escherichia Coli

Stress Response by Alternative Σ-factor, Rpoh, and Analysis of Posttranslational Modification of the Heat Shock Protein, Dnak, in Escherichia Coli

Date: May 2015
Creator: Martinez, Sarah N
Description: Bacteria have developed specialized responses that involve the expression of particular genes present in a given regulon. Sigma factors provide regulatory mechanisms to respond to stress by acting as transcriptional initiation factors. This work focuses on σ32 during oxidative stress in Escherichia coli. The differential response of key heat shock (HS) genes was investigated during HS and oxidative stress using qPCR techniques. While groEL and dnaJ experienced increases in transcriptional response to H2O2 (10 mM), HS (42°C), and paraquat (50 mM) exposure, the abundance of dnaK over the co-chaperones was apparent. It was hypothesized that DnaK undergoes oxidative modification by reactive carbonyls at its Lys-rich C-terminus, accounting for the differential response during oxidative stress. A σ32-mediated β-galactosidase reporter was devised to detect the activity of wild-type DnaK and DnaKV634X modified to lack the Lys-rich C-terminus. Under unstressed conditions and HS, σ32 was bound at the same rate in both strains. When subjected to H2O2, the WT DnaK strain produced significantly higher β-galactosidase than DnaKV634X (one-tailed Student’s t test p=0.000002, α=0.05) and approached the same level of output as the lacZ positive control. The β-galactosidase assay indicates that DnaK undergoes Lys modification in the WT strain, preventing the protein from binding ...
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Understanding Microbial Biodegradation of Environmental Contaminants

Understanding Microbial Biodegradation of Environmental Contaminants

Access: Use of this item is restricted to the UNT Community.
Date: May 2015
Creator: Vilo Muñoz, Claudia Andrea
Description: The accumulation of industrial contaminants in the natural environments have rapidly become a serious threat for human and animal life. Fortunately, there are microorganisms capable of degrading or transforming environmental contaminants. The present dissertation work aimed to understand the genomic basis of microbial degradation and resistance. The focus was the genomic study of the following bacteria: a) Pseudomonas fluorescens NCIMB 11764, a unique bacterium with specific enzymes that allow cyanide adaptation features. Potential cyanide degradation mechanisms found in this strain included nit1C cluster, and CNO complex. Potential cyanide tolerance genes found included cyanide insensitive oxidases, nitric oxide producing gene, and iron metabolism genes. b) Cupriavidus sp. strain SK-3 and strain SK-4. The genome of both bacteria presented the bph operon for polychlorinated biphenyl (PCB) degradation, but we found differences in the sequences of the genes. Those differences might indicate their preferences for different PCB substrates. c) Arsenic resistant bacterial communities observed in the Atacama Desert. Specific bacteria were found to thrive depending on the arsenic concentration. Examples were Bacteroidetes and Spirochaetes phyla whose proportions increased in the river with high arsenic concentrations. Also, DNA repair and replication metabolic functions seem to be necessary for resistance to arsenic contaminated environments. Our ...
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Acute Effects of the Antibiotic Streptomycin on Neural Network Activity and Pharmacological Responses

Acute Effects of the Antibiotic Streptomycin on Neural Network Activity and Pharmacological Responses

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Date: December 2014
Creator: Zeng, Wei Rong
Description: The purpose of this study is to find out that if antibiotic streptomycin decreases neuronal network activity or affects the pharmacological responses. The experiments in this study were conducted via MEA (multi-electrode array) technology which records neuronal activity from devices that have multiple small electrodes, serve as neural interfaces connecting neurons to electronic circuitry. The result of this study shows that streptomycin lowered the spike production of neuronal network, and also, sensitization was seen when neuronal network pre-exposed to streptomycin.
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Effects of a Water Conservation Education Program on Water Use in Single-family Homes in Dallas, Texas

Effects of a Water Conservation Education Program on Water Use in Single-family Homes in Dallas, Texas

Date: December 2014
Creator: Serna, Victoria Faubion
Description: The City of Dallas Environmental Education Initiative (EEI) is a hands-on, inquiry-based, K-12 water conservation education program that teaches students concepts about water and specific water conservation behaviors. Few descriptions and evaluations, especially quantitative in nature, of water conservation education programs have previously been conducted in the literature. This research measured the quantitative effects and impacts of the education program on water use in single-family homes in Dallas, Texas. A total of 2,122 students in 104 classrooms at three schools in the Dallas Independent School District received hands-on, inquiry-based water conservation education lessons and the average monthly water use (in gallons) in single-family homes was analyzed to measure whether or not there was a change in water use. The results showed that over a period of one calendar year the water use in the single-family homes within each school zone and throughout the entire research area in this study experienced a statistically significant decrease in water use of approximately 501 gallons per home per month (independent, t-test, p>0.001). Data from this research suggests that EEI is playing a role in decreasing the amount of water used for residential purposes. Additionally, this research demonstrates the use of a quantitative tool by ...
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Evidence for Multiple Functions of a Medicago Truncatula Transporter

Evidence for Multiple Functions of a Medicago Truncatula Transporter

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Date: December 2014
Creator: Huang, Ying-Sheng
Description: Legumes play an important role in agriculture as major food sources for humans and as feed for animals. Bioavailable nitrogen is a limiting nutrient for crop growth. Legumes are important because they can form a symbiotic relationship with soil bacteria called rhizobia that results in nitrogen-fixing root nodules. In this symbiosis, rhizobia provide nitrogen to the legumes and the legumes provide carbon sources to the rhizobia. The Medicago truncatula NPF1.7/NIP/LATD gene is essential for root nodule development and also for proper development of root architecture. Work in our lab on the MtNPF1.7/MtNIP/LATD gene has established that it encodes a nitrate transporter and strongly suggests it has another function. Mtnip-1/latd mutants have pleiotropic defects, which are only partially explained by defects in nitrate transport. MtNPF1.7/NIP/LATD is a member of the large and diverse NPF/NRT1(PTR) transporter family. NPF/NRT1(PTR) members have been shown to transport other compounds in addition to nitrate: nitrite, amino acids, di- and tri-peptides, dicarboxylates, auxin, abscisic acid and glucosinolates. In Arabidopsis thaliana, the AtNPF6.3/NRT1.1( CHL1) transporter was shown to transport auxin as well as nitrate. Atchl1 mutants have defects in root architecture, which may be explained by defects in auxin transport and/or nitrate sensing. Considering the pleiotropic phenotypes observed ...
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In Vitro Investigations of Antibiotic Influences on Nerve Cell Network Responses to Pharmacological Agents

In Vitro Investigations of Antibiotic Influences on Nerve Cell Network Responses to Pharmacological Agents

Date: December 2014
Creator: Sawant, Meera
Description: Neuronal networks, derived from mouse embryonic frontal cortex (FC) tissue grown on microelectrode arrays, were used to investigate effects of gentamicin pretreatment on pharmacological response to the L-type calcium channel blocker, verapamil. Gentamicin is a broad spectrum antibiotic used to control bacterial contamination in cell culture. The addition of gentamicin directly to medium affects the pharmacological and morphological properties of the cells in culture. A reproducible dose response curve to verapamil from untreated cultures was established and the mean EC50 was calculated to be 1.5 ± 0.5 μM (n=10). 40 μM bicuculline was added to some cell cultures to stabilize activity and verapamil dose response curves were performed in presence of bicuculline, EC50 1.4 ± 0.1 μM (n=9). Statistical analysis showed no significant difference in verapamil EC50s values obtained in presence of bicuculline and hence the data was combined and a standard verapamil EC50 was calculated as 1.4 ± 0.13 μM (n=19). This EC50 was then used to compare verapamil EC50s obtained from neuronal cell cultures with chronic and acute exposures to gentamicin. FC cultures (21- 38 days old) were found to be stable in presence of 2300 μM gentamicin. The recommended concentration of gentamicin for contamination control is 5uL ...
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Measuring Biomarkers From Dried Blood Spots Utilizing Bead-based Multiplex Technology

Measuring Biomarkers From Dried Blood Spots Utilizing Bead-based Multiplex Technology

Date: December 2014
Creator: Prado, Eric A
Description: Dried blood spots is an alternative method to collect blood samples from research subjects. However, little is known about how hemoglobin and hematocrit affect bead-based multiplex assay performance. The purpose of this study was to determine how bead-based multiplex assays perform when analyzing dried blood spot samples. A series of four experiments outline the study each with a specific purpose. A total of 167 subject samples were collected and 92 different biomarkers were measured. Median fluorescence intensity results show a positive correlation between filtered and non-filtered samples. Utilizing a smaller quantity of sample results in a positive correlation to a larger sample. Removal of hemoglobin from the dried blood spot sample does not increase detection or concentration of biomarkers. Of the 92 different biomarkers measured 56 were detectable in 100-75% of the attempted samples. We conclude that blood biomarkers can be detected using bead-based multiplex assays. In addition, it is possible to utilize a smaller quantity of sample while avoiding the use of the entire sample, and maintaining a correlation to the total sample. While our method of hemoglobin was efficient it also removed the biomarkers we wished to analyze. Thus, an alternative method is necessary to determine if removing ...
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Optimization of Novel Culturing and Testing Procedures for Acute Effects on Acartia Tonsa and Tisbe Biminiensis

Optimization of Novel Culturing and Testing Procedures for Acute Effects on Acartia Tonsa and Tisbe Biminiensis

Date: December 2014
Creator: Ussery, Erin J
Description: Copepods comprise an ecologically important role in freshwater and marine ecosystems, which is why they are often considered an important ecotoxicological model organism. The International Organization for Standardization’s (ISO) 14669 protocol is the only guideline for the determination of acute toxicity in three European marine copepod species: Acartia tonsa. The goal of this project was to assess the feasibility of establishing and maintaining cultures of Acartia tonsa, as well as to refine current culturing and egg separation methods. Initial culture methodology proved difficult for consistent production of eggs and collection of nauplii. The development of an airlift system for the separation of eggs from nauplii and adults, based on size, successfully increased the availability of eggs, nauplii and adults. The sensitivity and relative conditions of the copepod species was assessed by running a series of 48h acute toxicity tests with the reference toxicants 3,5-dichlorophenol, 4,4’-methylenebis(2,6-di-tert-butylphenol. The acute 48 hour median lethal dose concentration (LC50), the no observed effect concentration (NOEC), and the lowest observed effect concentration (LOEC) was analyzed for the three reference compounds for of A. tonsa.
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BK1 and DCD1 Act Synergistically in Subsidiary Cell Formation in Zea Mays

BK1 and DCD1 Act Synergistically in Subsidiary Cell Formation in Zea Mays

Date: August 2014
Creator: Malhotra, Divya
Description: Subsidiary mother cell (SMC) divisions during stomatal complex formation in Zea mays are asymmetric generating a small subsidiary cell (SC) and a larger epidermal cell. Mutants with a high number of abnormally shaped subsidiary cells include the brick1 (brk1) and discordia1 (dcd1) mutants. BRK1 is homologous to HSPC300, an ARP2/3 complex activator, and is involved in actin nucleation while DCD1 is a regulatory subunit of the PP2A phosphatase needed for microtubule generation (Frank and Smith, 2002; Wright et al. 2009). Possible causes of the abnormal SCs in brk1 mutants include a failure of the SMC nucleus to polarize in advance of mitosis, no actin patch, and transverse and/or no PPBs (Gallagher and Smith, 2000; Panteris et al 2006). The abnormal subsidiary mother cell division in dcd1 is due to correctly localized, but disorganized preprophase bands (PPBs; Wright et al. 2009). The observation that brk1 has defects in PPB formation and that the dcd1 phenotype is enhanced by the application of actin inhibitors led us to examine the dcd1; brk1 double mutant (Gallagher and Smith, 1999). We found that dcd1; brk1 double mutants demonstrate a higher percentage of aberrant SCs than the single mutants combined suggesting that these two mutations have ...
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Cloacal Microbiota of Captive-bred and Wild Attwater’s Prairie-chicken, Tympanuchus Cupido Attwateri

Cloacal Microbiota of Captive-bred and Wild Attwater’s Prairie-chicken, Tympanuchus Cupido Attwateri

Date: August 2014
Creator: Simon, Stephanie E.
Description: The Attwater’s prairie-chicken (Tympanuchus cupido attwateri; APC) is a species of grouse native to Texas coastal prairies and is on the critically endangered species list as a result of habitat destruction and overhunting. All of the current populations were captively bred and released into the wild. Survivorship for released APCs is very low, and individuals seldom survive to reproduce in the wild. One factor contributing to this may be an alteration in the gut microbiota as a result of captivity. Factors potentially influencing the gut microbial composition in captivity include antibiotic therapy, stress, and a predominantly commercially formulated diet. Recent studies have begun to shed light on the importance of the host microbial endosymbionts. Antibiotic administration, stress, diet, age, genotype and other factors have been shown to influence microbial populations in the gastrointestinal tracts of many different vertebrates. Sequencing of 16S rRNA gene amplicons on the Ion Torrent™ platform was used in this study to identify groups of bacteria in the cloacas as a surrogate for the gut microbiota in the APC. Antibiotic-treated and untreated birds, wild-hatched and captive-bred birds, and individuals sampled before and after release to the wild were examined. Significant differences were found between wild-hatched and captive ...
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