Genetics and Genomics Commons^™

Integral And Peripheral Association Of Proteins And Protein Complexes With Yersinia Pestis Inner And Outer Membranes, Rembert Pieper, Shih-Ting Huang, David J. Clark, Jeffrey M. Robinson, Hamid Alami, Prashanth P. Parmar, Moo-Jin Suh, Srilatha Kuntumalla, Christine L. Bunai, Robert D. Perry, Robert D. Fleischmann, Scott N. Peterson

Microbiology, Immunology, and Molecular Genetics Faculty Publications

Yersinia pestis proteins were sequentially extracted from crude membranes with a high salt buffer (2.5 M NaBr), an alkaline solution (180 mM Na2CO3, pH 11.3) and membrane denaturants (8 M urea, 2 M thiourea and 1% amidosulfobetaine-14). Separation of proteins by 2D gel electrophoresis was followed by identification of more than 600 gene products by MS. Data from differential 2D gel display experiments, comparing protein abundances in cytoplasmic, periplasmic and all three membrane fractions, were used to assign proteins found in the membrane fractions to three protein categories: (i) integral membrane proteins and peripheral membrane proteins with low solubility in …

One Rna Plays Three Roles To Provide Catalytic Activity To A Group I Intron Lacking An Endogenous Internal Guide Sequence, Nilesh Vaidya, Niles Lehman

Chemistry Faculty Publications and Presentations

Catalytic RNA molecules possess simultaneously a genotype and a phenotype. However, a single RNA genotype has the potential to adopt two or perhaps more distinct phenotypes as a result of differential folding and/or catalytic activity. Such multifunctionality would be particularly significant if the phenotypes were functionally inter-related in a common biochemical pathway. Here, this phenomenon is demonstrated by the ability of the Azoarcus group I ribozyme to function when its canonical internal guide sequence (GUG) has been removed from the 5? end of the molecule, and added back exogenously in trans. The presence of GUG triplets in noncovalent fragments of …

Microarray And Cdna Sequence Analysis Of Transcription During Nerve-Dependent Limb Regeneration, James R. Monaghan, Leonard G. Epp, Srikrishna Putta, Robert B. Page, John A. Walker, Chris K. Beachy, Wei Zhu, Gerald M. Pao, Inder M. Verma, Tony Hunter, Susan V. Bryant, David M. Gardiner, Tim T. Harkins, S. Randal Voss

Biology Faculty Publications

Background

Microarray analysis and 454 cDNA sequencing were used to investigate a centuries-old problem in regenerative biology: the basis of nerve-dependent limb regeneration in salamanders. Innervated (NR) and denervated (DL) forelimbs of Mexican axolotls were amputated and transcripts were sampled after 0, 5, and 14 days of regeneration.

Results

Considerable similarity was observed between NR and DL transcriptional programs at 5 and 14 days post amputation (dpa). Genes with extracellular functions that are critical to wound healing were upregulated while muscle-specific genes were downregulated. Thus, many processes that are regulated during early limb regeneration do not depend upon nerve-derived factors. …

High Resolution Single Molecule Optical Localization Of Multiple Fluorophores On Dna Origami Constructs Fluorophores On Dna Origami Constructs, Anuradha Rajulapati

Theses, Dissertations and Capstones

An ongoing challenge in the development of nanoelectronics and nanophotonics is the nondestructive, high-resolution localization in space of single molecules and multi-molecular assemblies. The apparent barrier to the use of optical microscopy at the sub-100 nm scale is the well known Abbe Limit, the diffraction limit to resolution. This laboratory has adapted a technique called Single-molecule high resolution imaging with photobleaching (SHRIMP). We have developed methods for utilizing SHRIMP for the determination of the separation of two fluorophores in single DNA origami constructs.

DNA Origami is extremely useful because it can address nanocomponents down to 2nm separation. It should be …

Characterization Of 24 Microsatellite Loci In Delta Smelt, Hypomesus Transpacificus, And Their Cross-Species Amplification In Two Other Smelt Species Of The Osmeridae Family, Kathleen M. Fisch, Jessica Lynn Petersen, Melinda R. Baerwald, John K. Pedroia, Bernie May

Department of Animal Science: Faculty Publications

We characterized 24 polymorphic tetranucleotide microsatellite loci for delta smelt (Hypomesus transpacificus) endemic to the San Francisco Bay Estuary, California, USA. Screening of samples (n = 30) yielded two to 26 alleles per locus with observed levels of heterozygosity ranging from 0.17 to 1.0. Only one locus deviated from Hardy–Weinberg equilibrium, suggesting these individuals originate from a single panmictic population. Linkage disequilibrium was found in two pairs of loci after excluding the locus out of Hardy–Weinberg equilibrium. Twenty-two primer pairs cross-amplified in wakasagi smelt (Hypomesus nipponensis), and 15 primer pairs cross-amplified in longfin smelt ( …

Letter From The Dean, Lalit Verma

Discovery, The Student Journal of Dale Bumpers College of Agricultural, Food and Life Sciences

No abstract provided.

Patterning Defects In Silkworm Embryos Analysed Through Cuticle Preparations, Amit Singh, Madhuri Kango-Singh, K. P. Gopinathan

Biology Faculty Publications

The mulberry silkworm, Bombyx mori, a holometabolous lepidopteran insect, has a metameric body plan. Due to its functional adaptation, B. mori presents some unique deviations in its pal/ern from the evolutionarily advanced dipteran insect, Drosophila melanogaster. Previous studies on mutant phenotype analysis in B. mori have been carried out in late stages of larval development. Here we employ, the cuticle preparation approach during embryonic development to study morphological landmarks associated with B. mori, Eri, another race a/silkworm, and pattern defects associated with Ekp mutant of B.mori. The homeotic mutant Ekp, generates ectopic abdominallegs, a feature …

Choosing The Right Path: Enhancement Of Biologically Relevant Sets Of Genes Or Proteins Using Pathway Structure, Reuben Thomas, Julia M. Gohlke, Geffrey F. Stopper, Frederick M. Parham, Christopher J. Portier

Biology Faculty Publications

A method is proposed that finds enriched pathways relevant to a studied condition using the measured molecular data and also the structural information of the pathway viewed as a network of nodes and edges. Tests are performed using simulated data and genomic data sets and the method is compared to two existing approaches. The analysis provided demonstrates the method proposed is very competitive with the current approaches and also provides biologically relevant results.

Site Directed Mutagensis Of Bacteriophage Hk639 And Identification Of Its Integration Site, Madhuri Jonnalagadda

Masters Theses & Specialist Projects

Bacteriophages affect bacterial evolution, pathogenesis and global nutrient cycling. They are also the most numerous and diverse group of biological entities on the planet [1, 2, 3, 4, 5, 6]. Members of the Lambda phage family share a similar genetic organization and control early gene expression by suppressing transcription, a process known as antitermination. Transcription antitermination in Lambda is mediated by a phage-encoded protein whereas in lambdoid phage HK022, antitermination is mediated by a phage-encoded RNA molecules. Recent results suggest that another bacteriophage called HK639 also appears to use RNA-mediated antitermination. To characterize this newly identified phage we generated site …

Evolutionary Genetics Of Tetrodotoxin (Ttx) Resistance In Snakes: Tracking A Feeding Adaptation From Populations Through Clades, Chris R. Feldman

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Understanding the nature of adaptive evolution has been the recent focus of research detailing the genetic basis of adaptation and theoretical work describing the mechanics of adaptive evolution. Nevertheless, key questions regarding the process of adaptive evolution remain. Ultimately, a detailed description of the ecological context, evolutionary history, and genetic basis of adaptations is required to advance our understanding of adaptive evolution. To address some of the contemporary issues surrounding adaptive evolution, I examine phenotypic and genotypic changes in a snake feeding adaptation.

Adaptations can arise through fixation of novel mutations or recruitment of existing variation. Some populations of the …

A Cell Biology Laboratory Exercise To Study Sub-Cellular Organelles In Drosophila, Meghana Tare, Amit Singh

Biology Faculty Publications

The fast-changing scenario of undergraduate education puts emphasis on introducing students to hands-on techniques as part of their laboratory courses. In order to cater to large numbers of students and the time constraints involved with undergraduate level laboratory courses, there is a need for development of experiments that are cost effective and can be completed in a defined time frame. We have devised a laboratory exercise for teaching cell biology using the Drosophila melanogaster model. Drosophila can be reared in a short period of time in a cost effective manner. We used Drosophila tissue to study the sub-cellular organization of …

Geographic Variation In Malarial Parasite Lineages In The Common Yellowthroat (Geothlypis Trichas), K. M. Pagenkopp, John Klicka, K. L. Durrant, J. C. Garvin, R. C. Fleischer

Ornithology Program (HRC)

Our current understanding of migration routes of many birds is limited and researchers have employed various methods to determine migratory patterns. Recently, parasites have been used to track migratory birds. The objective of this study was to determine whether haemosporidian parasite lineages detect significant geographic structure in common yellowthroats (Geothlypis trichas). We examined liver tissue or blood from 552 birds sampled from multiple locations throughout the continental United States, southern Canada, and the Bahamas. We found a 52.7% overall prevalence of haematozoan infection. We identified 86.1% of these infections to genus: 81% were Plasmodium; 5% were Haemoproteus …

Transcriptional Response Of Mexican Axolotls To Ambystoma Tigrinum Virus (Atv) Infection, Jennifer D. Cotter, Andrew Storfer, Robert B. Page, Christopher K. Beachy, S. Randal Voss

Biology Faculty Publications

Background

Very little is known about the immunological responses of amphibians to pathogens that are causing global population declines. We used a custom microarray gene chip to characterize gene expression responses of axolotls (Ambystoma mexicanum) to an emerging viral pathogen, Ambystoma tigrinum virus (ATV).

Result

At 0, 24, 72, and 144 hours post-infection, spleen and lung samples were removed for estimation of host mRNA abundance and viral load. A total of 158 up-regulated and 105 down-regulated genes were identified across all time points using statistical and fold level criteria. The presumptive functions of these genes suggest a robust …

Accelerated High Fidelity Prion Amplification Within And Across Prion Species Barriers, Kristi M. Green, Joaquín Castilla, Tanya S. Seward, Dana L. Napier, Jean E. Jewell, Claudio Soto, Glenn C. Telling

Microbiology, Immunology, and Molecular Genetics Faculty Publications

Experimental obstacles have impeded our ability to study prion transmission within and, more particularly, between species. Here, we used cervid prion protein expressed in brain extracts of transgenic mice, referred to as Tg(CerPrP), as a substrate for in vitro generation of chronic wasting disease (CWD) prions by protein misfolding cyclic amplification (PMCA). Characterization of this infectivity in Tg(CerPrP) mice demonstrated that serial PMCA resulted in the high fidelity amplification of CWD prions with apparently unaltered properties. Using similar methods to amplify mouse RML prions and characterize the resulting novel cervid prions, we show that serial PMCA abrogated a transmission barrier …

Cd5 Plays An Inhibitory Role In The Suppressive Function Of Murine Cd4(+) Cd25(+) T(Reg) Cells., Trivikram Dasu, Joseph E Qualls, Halide Tuna, Chander Raman, Donald A Cohen, Subbarao Bondada

Microbiology, Immunology, and Molecular Genetics Faculty Publications

A subset of CD4(+) T cells, the CD4(+) CD25(+) regulatory T (T(reg)) cells in the lymphoid organs and peripheral blood are known to possess suppressive function. Previous in vitro and in vivo studies have indicated that T cell receptor (TCR) signal is required for development of such 'natural regulatory (T(reg)) cells' and for activation of the effector function of CD4(+) CD25(+) regulatory T cells. CD5 is a cell surface molecule present on all T cells and a subtype of B lymphocytes, the B-1 cells, primarily localized to coelomic cavities, Peyer's patches, tonsils and spleen. CD5 acts as a negative regulator …

Cd5 Plays An Inhibitory Role In The Suppressive Function Of Murine Cd4+ Cd25+ TReg Cells, Trivikram Dasu, Joseph E. Qualls, Halide Tuna, Chander Raman, Donald A. Cohen, Subbarao Bondada

Microbiology, Immunology, and Molecular Genetics Faculty Publications

A subset of CD4⁺ T cells, the CD4⁺ CD25⁺ regulatory T (T_reg) cells in the lymphoid organs and peripheral blood are known to possess suppressive function. Previous in vitro and in vivo studies have indicated that T cell receptor (TCR) signal is required for development of such ‘natural regulatory (T_reg) cells’ and for activation of the effector function of CD4⁺ CD25⁺ regulatory T cells. CD5 is a cell surface molecule present on all T cells and a subtype of B lymphocytes, the B-1 cells, primarily localized to coelomic cavities, Peyer's patches, …

Expression Of Thor Does Not Increase Desiccation Resistance In Drosophila Melanogaster, Robert L. Kobey, Deborah K. Hoshizaki, Allen G. Gibbs

Undergraduate Research Opportunities Program (UROP)

Using microarray analysis of Drosophila melanogaster, the Gibbs lab has identified several hundred candidate genes that may be involved in desiccation resistance. One of these genes is Thor, an important downstream target of the TOR/insulin signaling pathway. Preliminary results confirm that Thor plays a role in desiccation resistance. Further research will be needed to verify these results and understand the mechanism by which Thor increases desiccation resistance. This research will also serve as a proof-of-principle for testing microarray-derived hypotheses.

A previous microarray analysis found evidence that down-regulation of protein synthesis might be a cellular response to desiccation through the up-regulation …

Construction Of A Thif Genetic Disruption In Bacillus Subtilis, Kathleen Bradley, Christine Pybus, Ronald Yasbin, Eduardo Robleto

Undergraduate Research Opportunities Program (UROP)

The goal of our research is to determine whether the level of transcription of a gene is correlated with the level of mutation in that gene. One factor involved in the mutability of a transcribed gene is the ability of the single stranded DNA to form secondary stem loop structures (SLS), in the wake of the transcription bubble, that contain unpaired mutable bases. We are interested in correlating the levels of mutation with transcription in the thiF gene, which is predicted by bioinformatic analysis to be highly mutable. To achieve this goal, Kathleen will first construct a non-polar thiF genetic …

Constructing An Argf- Strain Of Bacillus Subtilis, Allison Faucher, Christine Pybus, Ronald E. Yasbin, Eduardo A. Robleto

Undergraduate Research Opportunities Program (UROP)

The goal of our research is to determine whether the level of transcription of a gene is correlated with the level of mutation in that gene. One factor involved in the mutability of a transcribed gene is the ability of the single stranded DNA to form secondary stem loop structures (SLS), in the wake of the transcription bubble, that contain unpaired mutable bases. We are interested in correlating the levels of mutation with transcription in the argF gene, which is predicted by bioinformatic analysis to be highly mutable. To achieve this goal, Allison will first construct a non-polar argF genetic …

Decoding The Protein Interaction Network - An Approach Integrating Biology And Math, Ryan Huang, Lingkun Gu, Qingxi J. Shen

Undergraduate Research Opportunities Program (UROP)

The WRKY super family is known to play a major role during the plant stress response and development. My project focuses on the protein-protein interaction of an Oryzasativa (rice) transcription factor, OsWRKY71 which functions as the repressor of gibberellins signaling pathway. Previous literature revealed that OsWRKY71 can interact with itself or OsWRKY51 to form dimmers by using bimolecular fluorescence complementation (BiFC). To confirm this result, we use yeast two-hybrid system. As our data showed, OsWRKY71 seems to suppress the reporter gene expression of the conventional yeast two-hybrid system, so we use a modified yeast two-hybrid, Mating-based Split Ubiquitin System (MbSUS). …

Oxidative Stress-Mediated Anticancer Activity Of Novel Ahr Modulators Af & 5f203, Lancelot S. Mclean

Loma Linda University Electronic Theses, Dissertations & Projects

Estrogen receptor positive (ER+) breast cancer tends to respond to anti-estrogen agents such as Tamoxifen. Approximately 40% of ER+ breast cancer is resistant to these agents and those that initially respond often acquire resistance. Estrogen receptor negative (ER-) breast cancer remains largely unresponsive to these agents. It is therefore vital to discover drugs that are potent in both forms of breast cancer. Aminoflavone, (5-amino-2, 3-fluorophenyl)-6,8-difluoro-7-methyl-4H-l-benzopyran-4-one; AF; NSC 686288) and 5F203, (2-[-Amino-3-methy phenyl]-5-flurobenzothiazole) are novel anticancer candidate agents that display potent in vitro and in vivo anti-proliferative activity against select human tumor cells with a unique anticancer activity profile in the …

Novel Binding Domains Mediate Binding Of Hpv 16 E6 To Fadd And Procaspase 8, Sandy S. Tungteakkhun

Loma Linda University Electronic Theses, Dissertations & Projects

To evade the host response to infection, viruses have developed means to survive and propagate. HPV 16, a causative agent of cervical cancer and of some cases of oropharyngeal cancers, is one example. We have reported that the early viral protein E6 binds to proteins necessary for propagation of the apoptotic signal following receptor/ligand interactions, such as those mediated by FADD DED and procaspase 8 DED. E6 expression leads to the dose-dependent accelerated degradation of FADD and the protection of E6-expressing cells from Fas-induced apoptosis. Surprisingly, the splice isoforms of E6, E6_large and E6*, affect the stability of procaspase …

Mechanisms Of T Helper Cell Modulation In A Simulated Space Radiation Environment, Asma Rizvi

Loma Linda University Electronic Theses, Dissertations & Projects

Exposure to solar particle events (SPEs) poses the most serious threat to astronaut health. Due to this radiation environment, exploration of the solar system beyond Earth’s orbit entails many risks for the crew on these deep space missions. Depression or dysfunction of the immune system is a major concern due to its high radio-sensitivity; potential consequences include overwhelming infection, development of cancer, fibrosis, and poor wound healing. Astronauts are also exposed to low dose/low dose rate (LDR) radiation. A three month stay at the international space station exposes them to ~ 0.05 Gy of radiation.

C57BL/6 mice received LDR gamma-radiation …

Effect Of Thyroid Hormone Concentration On The Transcriptional Response Underlying Induced Metamorphosis In The Mexican Axolotl (Ambystoma), Robert B. Page, Stephen R. Voss, Amy K. Samuels, Jeramiah J. Smith, Srikrishna Putta, Christopher K. Beachy

Biology Faculty Publications

Background

Thyroid hormones (TH) induce gene expression programs that orchestrate amphibian metamorphosis. In contrast to anurans, many salamanders do not undergo metamorphosis in nature. However, they can be induced to undergo metamorphosis via exposure to thyroxine (T₄). We induced metamorphosis in juvenile Mexican axolotls (Ambystoma mexicanum) using 5 and 50 nM T₄, collected epidermal tissue from the head at four time points (Days 0, 2, 12, 28), and used microarray analysis to quantify mRNA abundances.

Results

Individuals reared in the higher T₄ concentration initiated morphological and transcriptional changes earlier and completed metamorphosis by …

Historic Genetic Structuring And Paraphyly Within The Great-Tailed Grackle, Jeffrey M. Dacosta, Walter Wehtje, John Klicka

Ornithology Program (HRC)

The Great-tailed Grackle (Quiscalus mexicanus) and Boat-tailed Grackle (Q. major) are sister species that have expanded their ranges during historical times. This expansion has created an area of sympatry between these species in Texas and Louisiana, and between distinctive Great-tailed Grackle subspecies in the southwestern United States and northern Mexico. We investigated the evolutionary histories of both species using mitochondrial DNA sequence data and modern phylogenetic methods. Our results reveal genetic structure within Great-tailed, but not Boat-tailed Grackles. Great-tailed Grackles are separated into two clades, but range expansion in the north has led to secondary contact …

Optimizing Qpcr For The Quantification Of Periodontal Pathogens In A Complex Plaque Biofilm, Sreenatha S. Kirakodu, M. Govindaswami, Michael John Novak, Jeffrey L. Ebersole, Karen F. Novak

Center for Oral Health Research Faculty Publications

Quantitative PCR (qPCR) has recently been used to quantify microorganisms in complex communities, including dental plaque biofilms. However, there is variability in the qPCR protocols being used. This study was designed to evaluate the validity of two of these variables with the intent of developing a more standardized qPCR protocol. The two variables evaluated were (1) the use of DNA content versus actual cell counts to estimate bacterial numbers in mixed plaque samples and (2) the effectiveness of three different universal primers versus species specific primers in amplifying specific target pathogens in these samples. Results lead to the development of …

Molecular Evolution Of The Cytochrome C Oxidase Subunit 5a Gene In Primates, Monica Uddin, Juan C. Opazo, Derek E. Wildman, Chet C. Sherwood, Patrick R. Hof, Morris Goodman, Lawrence I. Grossman

Wayne State University Associated BioMed Central Scholarship

Abstract

Background

Many electron transport chain (ETC) genes show accelerated rates of nonsynonymous nucleotide substitutions in anthropoid primate lineages, yet in non-anthropoid lineages the ETC proteins are typically highly conserved. Here, we test the hypothesis that COX5A, the ETC gene that encodes cytochrome c oxidase subunit 5A, shows a pattern of anthropoid-specific adaptive evolution, and investigate the distribution of this protein in catarrhine brains.

Results

In a dataset comprising 29 vertebrate taxa, including representatives from all major groups of primates, there is nearly 100% conservation of the COX5A amino acid sequence among extant, non-anthropoid placental mammals. The most recent common …

1, Structural And Functional Studies Of Human Replication Protein A; 2 Dna Damage Responses And Dna Repair Defects In Laminopathy-Based Premature Aging., Yiyong Liu

Electronic Theses and Dissertations

The genome of mammalian cells is under constant attack from DNA-damaging agents. To maintain genomic integrity, cells activate an array of pathways primarily consisting of DNA repair and DNA damage checkpoints. Human replication protein A (RPA), a single-stranded DNA (ssDNA) binding protein, is essential for almost all DNA metabolic pathways. However, the role of RPA in nucleotide excision repair (NER), a DNA repair pathway for removing bulky DNA lesions, remains elusive. In this study, the binding of RPA to a battery of well-defined ssDNA substrates has been systematically examined using fluorescence spectroscopy. The results showed that RPA has a lower …

Mechanism Of Transcriptional Regulation Of C-Reactive Protein Gene Expression., Bhavya Voleti

Electronic Theses and Dissertations

C-reactive protein (CRP) is an acute phase protein produced by hepatocytes whose serum concentration increases in inflammatory conditions including cardiovascular complications. Statins that are used in the treatment of cardiovascular diseases to reduce cholesterol also lower serum CRP levels. In human hepatoma Hep3B cells, CRP is induced in response to cytokines IL-6 and IL-1β. The objective of the study was to determine the mechanism of regulation of CRP gene expression in Hep3B cells in response to cytokines and to determine the effect of statins on CRP expression. Key findings of our research were: 1. IL-1β-activated NF-κB p50/p65 acted synergistically with …

Mechanisms Of Covalent Self-Assembly Of The Azoarcus Ribozyme From Four Fragment Oligonucleotides, Will E. Draper, Eric John Hayden, Niles Lehman

Chemistry Faculty Publications and Presentations

RNA oligomers of length 40–60 nt can self-assemble into covalent versions of the Azoarcus group I intron ribozyme. This process requires a series of recombination reactions in which the internal guide sequence of a nascent catalytic complex makes specific interactions with a complement triplet, CAU, in the oligomers. However, if the CAU were mutated, promiscuous self-assembly may be possible, lessening the dependence on a particular set of oligomer sequences. Here, we assayed whether oligomers containing mutations in the CAU triplet could still self-construct Azoarcus ribozymes. The mutations CAC, CAG, CUU and GAU all inhibited self-assembly to some degree, but did …