A novel luminescence resonance energy transfer (LRET) nanocircuit assay involving a donor and two acceptors in tandem was developed to study the dynamic interaction of skeletal muscle contraction proteins. The donor transmits energy relayed to the acceptors distinguishing myosin subfragment-1 (S1) lever arm orientations. The last acceptor allows the detection of S1's bound near or in between troponin complexes on the thin filament. Additionally, calcium related changes between troponin T and myosin were detected. Based on this data, the troponin complex situated every 7 actin monomers, hinders adjacently bound myosins to complete their power stroke; whereas myosins bound in between troponin complexes undergo complete power strokes.

Superior swimming stamina in adult fish is presumably passed on to their offspring, but the ontogeny of the appearance of superior stamina and the requisite enhanced cardio-respiratory support for locomotion in larval fishes has not been determined. Is the expression of the suite of parental traits enabling superior swimming stamina in their offspring dependent upon their achieving juvenile/adult morphology, or does it appear earlier in their larvae? To answer this, adults were classified into three groups based on swimming stamina, followed by measurement of length, mass, and width. Larval offspring from the two parental groups -high stamina larvae (HSL) and low stamina larvae (LSL)- were reared at 27°C in aerated water (21% O2). Routine and active heart rate, routine and active mass specific oxygen consumption were recorded through 21dpf, and cost of transport (COT) and factorial aerobic scope were derived from oxygen consumption measurements. Routine heart rate at 2dpf of LSL was 164 ± 1 b·min-1, compared to only 125 ± 2 b·min-1 for HSL. Routine heart rate subsequently peaked at 203 ± 1 b·min-1 at 5dpf in the HSL group, compared to 207 ± 1 b·min-1, at 4dpf in the LSP larvae. Active heart rate at 5 dpf of …

A bacterial challenge assay was developed utilizing the earthworm, Lumbricus terrestris, in order to assess potential immunotoxic effects from exposure to specific polychlorinated biphenyl congeners. Earthworms were inoculated with Aeromonous hydrophila, establishing a 10-day LD50. In vitro assays for effects of PCBs on phagocytosis agreed with mammalian studies, demonstrating potent suppression of phagocytosis by the non-coplanar PCB congener 138 and no suppression by the coplanar congener 126. However, when the effects of the two PCB congeners were evaluated for suppression of resistance to a whole animal infection challenge assay, coplanar PCB 126 decreased the ability of L. terrestris to withstand infection while non-coplanar PCB 138 did not.

Large amounts of industrial fertilizers are used to maximize crop yields. Unfortunately, they are not completely consumed by plants; consequently, this leads to soil pollution and negative effects on aquatic systems. An alternative to industrial fertilizers can be found in legume plants that provide a nitrogen source that is not harmful for the environment. Legume plants, through their symbiosis with soil bacteria called rhizobia, are able to reduce atmospheric nitrogen into ammonia, a biological nitrogen source. Establishment of the symbiosis requires communication on the molecular level between the two symbionts, which leads to changes on the cellular level and ultimately results in nitrogen-fixing nodule development. Inside the nodules hypoxic environment, the bacterial enzyme nitrogenase reduces atmospheric nitrogen to ammonia. Medicago truncatula is the model legume plant that is used to study symbiosis with mycorrhiza and with the bacteria Sinorhizobium meliloti. The focus of this work is the M. truncatula nodulation mutant nip (numerous infections and polyphenolics). The NIP gene plays a role in the formation and differentiation of nodules, and development of lateral roots. Studying this mutant will contribute knowledge to understanding the plant response to infection and how the invasion by rhizobia is regulated. Previous genetic mapping placed NIP …

In order to assess the enhancement of seedling survival and growth during drought conditions, five-hundred bare-root seedlings each of Shumard oak (Quercus shumardii Buckl.) and green ash (Fraxinus pennsylvanica Marsh.) were planted each with four soil amendments at a Wildlife Management Area in Lewisville, Texas. The treatments were a mycorrhizal inoculant, mulch fabric, and two superabsorbent gels (TerraSorb® and DRiWATER®). Survival and growth measurements were assessed periodically for two years. Research was conducted on vegetation, soil, and site history for baseline data. Both superabsorbent gels gave significant results for Shumard oak survival, and one increased green ash diameter. For overall growth, significant results were found among DRiWATER®, mycorrhizae, and mulch treatments.

The rapid development of once-rural landscapes often produces detrimental effects on surface water quality entering local reservoirs through vulnerable stream channels. This study presents a methodology that incorporates geographic information systems (GIS) and remote sensing techniques for the creation of a stream corridor evaluation mechanism, coined the water quality corridor management (WQCM) model. Specifically, the study focuses on determining the viability of the WQCM model in assessing the stream corridor conditions within a northern Denton County pilot study region. These results will aid in the prediction and evaluation of the quality of stream water entering reservoirs that serve as the primary drinking water source for local municipalities.

The study of root nodule symbiosis between nitrogen-fixing bacteria and leguminous plant species is important because of the ability to supplement fixed nitrogen fertilizers and increase plant growth in poor soils. Our group has isolated a mutant called nip in the model legume Medicago truncatula that is defective in nodule symbiosis. The nip mutant (numerous infections with polyphenolics) becomes infected by Sinorhizobium meliloti but then accumulates polyphenolic defense compounds in the nodule and fails to progress to a stage where nitrogen fixation can occur. Analysis of the transcriptome of nip roots prior to inoculation with rhizobia was undertaken using Affymetric Medicago Genome Array microarrays. The total RNA of 5-day old uninoculated seedlings was analyzed in triplicate to screen for the NIP gene based on downregulated transcript levels in the mutant as compared to wild type. Further microarray data was generated from 10 days post inoculation (dpi) nip and wild type plants. Analysis of the most highly downregulated transcripts revealed that the NIP gene was not identifiable based on transcript level. Putative gene function was assigned to transcripts with altered expression patterns in order to characterize the nip mutation phenotypically as inferred from the transcriptome. Functional analysis revealed a large number …

Commonly, when it is desirable to replace an essential gene with an allelic series of mutated genes, or genes with altered expression patterns, the complementing constructs are introduced into heterozygous plants, followed by the selection of homozygous null segregants. To overcome this laborious and time-consuming step, the newly developed two-component system utilizes a site-specific recombinase to excise a wild-type copy of the gene of interest from transformed tissues. In the first component (the first vector), a wild-type version of the gene is placed between target sequences recognized by FLP recombinase from the yeast 2 μm plasmid. This construct is transformed into a plant heterozygous for a null mutation at the endogenous locus, and progeny plants carrying the excisable complementing gene and segregating homozygous knockout at the endogenous locus are selected. The second component (the second vector) carries the experimental gene along with the FLP gene. When this construct is introduced, FLP recombinase excises the complementing gene, leaving the experimental gene as the only functional copy. The FLP gene is driven by an egg apparatus specific enhancer (EASE) to ensure excision of the complementing cDNA in the egg cell and zygote following floral-dip transformation. The utility of this system is being …

The chicken ductus arteriosi (DA) are two embryonic blood vessels that shunt blood away from the non-ventilated lungs and towards the body and chorioallantoic membrane. I show that prostaglandins have a diminished role in maintaining chicken DA patency and nitric oxide inhibits oxygen induced contraction of the day 19 proximal DA in a time dependent manner. The pathways governing oxygen induced contraction in the chicken DA are similar to those found in mammals and include contributions from ROS, Kv channels, L-type Ca2+ channels, and the Rho kinase pathway. Longer exposure to high oxygen generates increased oxygen induced constriction of the day 19 DA that may be mediated through the Rho kinase pathway.

Raptor rehabilitation has become commonplace globally, yet studies on the survival and adaptation of great horned owls (Bubo virginianus) after release has been neglected to an appreciable extent. The primary objective of this study is to provide quantitative data on the success of rehabilitated and released great horned owls in the North Texas region. Owls (N=12) were rehabilitated and released onto the Ray Roberts Greenbelt Corridor in Denton County, Texas, and monitored using radio telemetry to evaluate home range (November 2002 - February 2005). With approximately 75% of the birds released for this study surviving until transmitter battery failure, it is believed that the rehabilitation process was successful for these birds.

The Medicago truncatula NIP gene is located on M. truncatula Linkage Group 1. Informative recombinants showed crossovers that localize the NIP gene between markers 146O17 and 23C16D. Marker 164N9 co-segregates with the NIP gene, and the location of marker 164N9 is between markers 146O17 and 23C16D. Based upon data from the Medicago genome sequencing project, a subset of the model legume Medicago truncatula bacterial artificial chromosomes (BACs) were used to create a physical map on the DNA in this genetic internal. BACs near the potential NIP gene location near marker 164N9 were identified, and used in experiments to predict the physical map by a BAC-by-BAC strategy. Using marker 164N9 as a center point, and chromosome walking outward, the physical map toward markers 146O17 and 23C16D was built. The chromosome walk consisted of a virtual walk, made with existing sequence of BACs from the Medicago genome project, hybridizations to filters containing BAC DNA, and PCR reactions to confirm that predicted overlapping BACs contained DNA that yielded similar PCR products. In addition, the primers which are made for physical mapping via PCR could be good genetic markers helpful in discovering the location of the NIP gene. As a result of efforts repotted …

This thesis describes the development of an optimized method for culturing dissociated, monolayer neuronal networks from murine frontal cortex and midbrain. It is presented as a guidebook for use by cell culture specialists and laboratory personnel who require updated and complete procedures for use with microelectrode array (MEA) recording technology. Specific cell culture protocols, contamination prevention and control, as well common problems encountered within the cell culture facility, are discussed. This volume offers value and utility to the rapidly expanding fields of MEA recording and neuronal cell culture. Due to increasing interest in determining the mechanisms underlying Parkinson's disease, the newly developed procedures for mesencephalon isolation and culture on MEAs are an important research contribution.