Molecular Biology Commons^™

Detection Of Insulin Receptor, Epidermal Growth Factor Receptor, And Interleukin-6 On Individual Mouse Embryos By Immuno-Polymerase Chain Reaction, Kun Xu

Electronic Theses and Dissertations

Three series of experiments were conducted to: 1) optimize the conditions for the production of pUC19 plasmid and of biotinylated pUC19 fragments; 2) optimize the conditions for the production of protein A-streptavidin chimera (chimeric protein); and 3) detect soluble antigens [bovine serum albumin (BSA) and interleukin-6 (IL-6)] and membrane-bound antigens [insulin receptor and epidermal growth factor receptor (EGFr)] of mouse embryos by immuno-polymerase chain reaction (I-PCR). The first experimental series, which included bacterial culture, chimeric protein purification, and chimeric protein functional experiments, was performed to investigate the effects of IPTG (isopropyl-β-D-thiogalactopyranoside) induction time and temperature, bacterial culture medium, and protein …

Cold-Temperature Adaptation Of Muscle Creatine Kinase From An Antartic Teleost (Chaenocephalus Aceratus), Paul Winnard Jr.

Electronic Theses and Dissertations

The white muscle of Chuenocephulus aceratus, an Antarctic teleost of the Channicthyidae family, has a compromised glycoiytic capacity and this fish cannot depend on glycolysis for rapid ATP generation For C. aceratus, creatine kinase (CK) and phosphocreatine (PCr) reserves comprise the metabolic pathway that may supplement and overcome this deficiency in energy transduction. Two conditions, low glycolytic capacity and evolution in a chronically cold habitat (-1.86°C), give us reason to believe that C. aceratus muscle CK (MMCK) has been subjected to strong selective pressure. Thus, the hypothesis of this thesis is that MMCK fiom C. aceratus white muscle …

Quinic Acid-Mediated Induction Of Hypovirulence And A Hypovirulence-Associated Double-Stranded Rna In Rhizoctonia Solani, Chunyu Liu

Electronic Theses and Dissertations

This study is a part of a project focused on the relationship between dsRNA and hypovirulence in R. solani. Hypovirulence refers to a condition in which a pathogen has a drastically reduced ability to cause disease. Rhizoctonia solani is a soil-borne pathogen causing diseases in numerous plants. Phenyl acetic acid (PAA), a metabolite of phenylalanine, causes Rhizoctonia disease symptoms on potato in the absence of the pathogen itself. The amount of PAA produced by a hypovirulent isolate is 10% of that produced by virulent isolates. A 3.6 kb dsRNA (M2) has been shown to be associated with hypovirulence in …

Interactions Among Murine Cytomegalovirus Us22 Family Gene Products That Influence Viral Pathogenesis, Zaruhi Karabekian

Theses and Dissertations in Biomedical Sciences

Cytomegalovirus (CMV) is a complex, ubiquitous herpesvirus that is characterized by acute, chronic, and latent infections. Monocytes-macrophages are the key target cell type involved in pathogenesis, which is most effectively studied using the murine model of CMV infection. Previously three murine CMV (MCMV) genes (M139, M140, and M141) were identified to regulate viral expression in cultured macrophages and in mice. These genes are members of the US22 gene family with respect to HCMV homology. There is no function assigned to the proteins encoded by these genes. However, deletion of M139, M140, and M141 significantly curtails growth of MCMV in macrophages …

Positive Regulation Of Pka On Human Gonadotropin-Releasing Hormone (Hgnrh) Gene Expression In Human Placental Jeg-3 Cells, Zhaoyang Wen

Chemistry & Biochemistry Theses & Dissertations

Using the human placental choriocarcinoma JEG-3 cell line as an in vitro human placental model, we studied the mechanisms of the PKA positive regulation of the hGnRH gene expression in the human placenta. Studies in JEG3 cells showed that through the PKA catalytic subunit a, human GnRH upstream promoter activity was stimulated by PKA signaling pathway in a cAMP dependent mechanism. The sequence between —202 (Afl II) and —554 (BamH I) base pair in the human GnRH upstream promoter region appeared to be responsible for the PKA positive regulation of the gene expression. Furthermore, Western blot analysis demonstrated the involvement …

Regulation Of The Hcmv Ul98 Promoter By Cooperation Of The Cyclic Amp Response Element And Gamma Interferon Response Element, Tabmitha Patricia Young Jervey

Theses and Dissertations in Biomedical Sciences

The HCMV UL98 early alkaline exonuclease gene promoter was examined to determine the DNA sequences as well as viral and/or cellular proteins functional in the regulation of this early gene. To assess promoter activation, UL98 promoter sequences were first cloned upstream of the chloramphenicol acetyltransferase (CAT) reporter gene and cotransfected with expression plasmids which express the HCMV major immediate early (IE) proteins 1E72 and 1E86. To more specifically determine the importance of individual cis-acting elements in UL98 promoter activation, the promoter region underwent mutagenesis to delete or alter sequences. The variant promoters were again cloned into a reporter-CAT construct and …

Detection Of Aneuploidy For Chromosomes 7 And 8 Using Fluorescence In Situ Hybridization In Patients With Aplastic Anemia And Sequencing Of The Mitotic Checkpoint Gene Hbub1, Laura Jane Aridgides

Theses and Dissertations in Biomedical Sciences

Aplastic anemia (AA) is characterized by complete bone marrow failure. Progression to myelodysplastic syndromes (MDS) and acute nonlymphocytic leukemia (ANLL) occurs frequently. At the time of transformation, cytogenetic abnormalities are common. Detection of cytogenetic abnormalities prior to leukemic transformation may indicate future disease progression. Karyotype analysis is the current method of choice to evaluate chromosome aberrations. However, fluorescence in situ hybridization (FISH) is more sensitive in detecting these abnormalities.

hBUB1, a mitotic spindle checkpoint gene, was shown to be mutated in two colorectal cancer cell lines with high levels of aneuploidy (Cahill, et al., 1998). Although theoretically possible, conclusive …

Molecular Characterization Of Cultured Perkinsus Marinus Isolates, Gwynne D. Brown

Dissertations, Theses, and Masters Projects

Perkinsus marinus is the causative agent of the oyster disease Dermo in the eastern oyster, Crassostrea virginica. In vitro propagation of the parasite has led to the establishment of multiple isolates by several investigators. Little work, however, has been done to characterize different isolates. In this study multiple isolates were examined for genetic and biochemical diversity. at two loci, the ITS region and the ATAN region, there was as much intra-isolate genetic variation among DNA sequences of some isolates as there was inter-isolate variation. Variation was also observed at a third loci, a subtilisin-like serine protease gene. This is the …

Adrenal Expression Of Cyp17 And Hsd3b1 Mrna In Peromyscus Maniculatus Bairdii, Brian Thomas Lovitt

Dissertations, Theses, and Masters Projects

No abstract provided.

Characterization Of The Human Cytomegalovirus Ul75 Late Gene Promoter, Bernard J. P. Mcwatters

Theses and Dissertations in Biomedical Sciences

Gene expression during productive infection by the human cytomegalovirus (HCMV) occurs in an ordered and sequential manner, beginning with immediate early (IE), then early (E) and finally late (L) gene expression. Significant work has addressed the regulation of IE and E gene expression while relatively little work has addressed the control of late gene expression. In order to further address HCMV late gene expression, the promoter of the HCMV UL75 (glycoprotein H, gH) late gene was characterized. The data obtained in this study were combined with observations made in two other studies that have addressed HCMV late gene expression to …

Characterization Of The Biological Activities Of Recombinant Fusion Protein Green Fluorescent Protein/Human Zona Pellucida Protein 3 (Gfp/Hzp3), Zhiyong Lin

Theses and Dissertations in Biomedical Sciences

Despite numerous reports indicating the successful production of bioactive recombinant ZP3, no report has shown the rhZP3 having direct binding activity with human sperm. Recombinant ZP3 generated from our previous study displayed binding activity with human sperm through indirect evidence from hemizona assay (HZA).

This present study focused on the production of recombinant ZP3 with direct binding activity with human sperm. Through the application of a pEGFP expression vector, fusion protein GFP/ZP3 was successfully generated and expressed. The expression of GFP/ZP3 was evidenced by RT-PCR and western blot. The fusion protein was partially purified by Ni-NTA affinity column from cell …