Biotechnology Commons^™

Antisense Oligonucleotides: Treatment Strategies And Cellular Internalization, Colton M. Miller, Ed Harris

Department of Biochemistry: Faculty Publications

The clinical application of antisense oligonucleotides (ASOs) is becoming more of a reality as several drugs have been approved for the treatment of human disorders and many others are in various phases in development and clinical trials. ASOs are short DNA/RNA oligos which are heavily modified to increase their stability in biological fluids and retain the properties of creating RNA-RNA and DNA-RNA duplexes that knock-down or correct genetic expression. This review outlines several strategies that ASOs utilize for the treatment of various congenital diseases and syndromes that develop with aging. In addition, we discuss some of the mechanisms for specific …

Pch1 Integrates Circadian And Light- Signaling Pathways To Control Photoperiod-Responsive Growth In Arabidopsis, He Huang, Chan Yul Yoo, Rebecca Bindbeutel, Jessica Goldsworthy, Allison Tielking, Sophie Alvarez, Michael J. Naldrett, Bradley S. Evans, Meng Chen, Dmitri A. Nusinow

Nebraska Center for Biotechnology: Faculty and Staff Publications

Plants react to seasonal change in day length through altering physiology and development. Factors that function to harmonize growth with photoperiod are poorly understood. Here we characterize a new protein that associates with both circadian clock and photoreceptor components, named PHOTOPERIODIC CONTROL OF HYPOCOTYL1 (PCH1). pch1 seedlings have overly elongated hypocotyls specifically under short days while constitutive expression of PCH1 shortens hypocotyls independent of day length. PCH1 peaks at dusk, binds phytochrome B (phyB) in a red light-dependent manner, and co-localizes with phyB into photobodies. PCH1 is necessary and sufficient to promote the biogenesis of large photobodies to maintain an …

Vo-Ohpic Treatment Reduces Cardiac Remodeling In Doxorubicin-Induced Cardiomyopathy, Taylor Johnson

Electronic Theses and Dissertations

Doxorubicin (Doxo) is one of multiple anthracycline drugs used to effectively treat various forms of cancer. Unfortunately, Doxo treatment, as a side effect, induces cardiomyopathy and subsequent heart failure. We have previously demonstrated that transplanted embryonic stem (ES) cells and their conditioned medium (CM) modulate the PTEN pathway and reduce apoptosis, fibrosis and hypertrophy in a Doxo induced cardiomyopathy (DIC) model. However, mechanisms of inhibited apoptosis mediated through PTEN pathway are completely unknown. Therefore, we used VO-OHpic (VO), a potent PTEN inhibitor to understand the mechanism of apoptosis as well as its effect on cardiac remodeling in DIC. Animals were …

The Role Of Type-I Interferon In Limiting Spread And Killing Of An Oncolytic Rna Virus In Prostate Cells, Kritika Kedarinath

Electronic Theses and Dissertations

Prostate cancer is the second most prevalent cancer amongst men and there is an urgent need to address viable therapeutic options for its treatment. Development of viruses which target and kill cancer cells has gained momentum due to the first FDA approved oncolytic virus for treating human cancer patients. Our previous work with the RNA virus, Parainfluenza Virus 5 (PIV5), has led to the generation of mutants that are potential candidates for oncolytic viruses: 1) the hyperfusogenic (P/V/F) mutant has a mutated P/V and fusion gene which activates anti-viral responses and causes massive cell-cell fusion respectively, and 2) the Leader …

Malondialdehyde (Mda) And Glutathione Peroxidase (Gpx) Are Elevated In Crohns Disease-Associated With Mycobacterium Avium Subspecies Paratuberculosis (Map), Ahmad Qasem

Electronic Theses and Dissertations

Inflamed tissue in Crohn’s disease (CD) are continuously producing toxic oxygen metabolites leading to cellular injury and apoptosis. Here, we are evaluating the role of Mycobacterium avium subspecies paratuberculosis (MAP) in oxidative stress in CD by evaluation of lipid peroxidation and antioxidant defense activity. Specifically, we measured malondialdehyde (MDA) level and selenium-dependent glutathione peroxidase (GPx) activity in the plasma from patients and cattle infected with MAP. The level of MAP antibodies in bovine sera was determined by IDEXX kit whereas detection of MAP DNA was performed by IS900-based nPCR. A total of 42 cattle (21 infected with MAP and 21 …

Efficiency Of The Nuclease I-Scei In Excising Selectable Marker Genes From The Plant Genome, Elliot E. Pruett, Soumen Nandy, Vibha Srivastava

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

Gene stacking is a method used in biotechnology by which multiple genes can be placed at a single genomic site, thereby simplifying plant breeding. In this approach, DNA nucleases are used for excising selectable marker genes (SMG), which are the unneeded components of transgenic plants. The goal of this project is to evaluate the effectiveness of the nuclease I-SceI in excising DNA in plants. Specifically, this study tests heat-inducible I-SceI through the use of a heat-shock promoter (HS) in order to control SMG excision by heat application. The DNA plasmid containing a visual marker gene flanked by I-SceI target sites …

A Review Of Natural And Engineered Enzymes Involved In Bioethanol Production, Ines Cuesta Urena

Graduate Student Theses, Dissertations, & Professional Papers

Alternative petroleum-derived fuels, such as biofuels, is another form to decrease the dependence of non-renewable energy. The most promising alternative energy is cellulosic ethanol because of the abundance of cellulose and the overall lack of concern for the food-versus-fuel dilemma.

In order to produce ethanol from cellulosic materials, pretreatment is required to “open” the lignocellulosic matrix and make cellulose more susceptible to enzymatic degradation. Enzymatic hydrolysis of lignocellulose is an important area of research due to the absence of negative effects in downstream processes in comparison with acid hydrolysis. Both natural enzymes and engineered enzymes can be used in the …

The Cytotoxic Synergy Of Nanosecond Electric Pulses And Low Temperature Leads To Apoptosis, Claudia Muratori, Andrei G. Pakhomov, Elena C. Gianulis, Sarah D. Jensen, Olga N. Pakhomova

Bioelectrics Publications

Electroporation by nanosecond electric pulses (nsEP) is an emerging modality for tumor ablation. Here we show the efficient induction of apoptosis even by a non-toxic nsEP exposure when it is followed by a 30-min chilling on ice. This chilling itself had no impact on the survival of U-937 or HPAF-II cells, but caused more than 75% lethality in nsEP-treated cells (300 ns, 1.8-7 kV/cm, 50-700 pulses). The cell death was largely delayed by 5-23 hr and was accompanied by a 5-fold activation of caspase 3/7 (compared to nsEP without chilling) and more than 60% cleavage of poly-ADP ribose polymerase (compared …

Inactivation Of Acanthamoeba Spp. And Other Ocular Pathogens By Application Of Cold Atmospheric Gas Plasma, Wayne Heaselgrave, Gilbert Shama, Peter W. Andrew, Michael G. Kong

Bioelectrics Publications

Currently there are estimated to be approximately 3.7 million contact lens wearers in the United Kingdom and 39.2 million in North America. Contact lens wear is a major risk factor for developing an infection of the cornea known as keratitis due to poor lens hygiene practices. While there is an international standard for testing disinfection methods against bacteria and fungi (ISO 14729), no such guidelines exist for the protozoan Acanthamoeba, which causes a potentially blinding keratitis most commonly seen in contact lens wearers, and as a result, many commercially available disinfecting solutions show incomplete disinfection after 6 and 24 …

Role Of Apoptotic Hepatocytes In Hcv Dissemination: Regulation By Acetaldehyde, Murali Ganesan, Sathish Kumar Natarajan, Jinjin Zhang, Justin L. Mott, Larisa I. Poluektova, Benita L. Mcvicker, Kusum K. Kharbanda, Dean J. Tuma, Natalie A. Osna

Department of Biochemistry: Faculty Publications

Alcohol consumption exacerbates hepatitis C virus (HCV) pathogenesis and promotes disease progression, although the mechanisms are not quite clear. We have previously observed that acetaldehyde (Ach) continuously produced by the acetaldehyde-generating system (AGS), temporarily enhanced HCV RNA levels, followed by a decrease to normal or lower levels, which corresponded to apoptosis induction. Here, we studied whether Ach-induced apoptosis caused depletion of HCV-infected cells and what role apoptotic bodies (AB) play in HCV-alcohol crosstalk. In liver cells exposed to AGS, we observed the induction of miR-122 and miR-34a. As miR-34a has been associated with apoptotic signaling and miR-122 with HCV replication, …

Inhibition Of Protein Ubiquitination By Paraquat And 1-Methyl-4-Phenylpyridinium Impairs Ubiquitin-Dependent Protein Degradation Pathways, Juliana Navarro-Yepes, Anandhan Annadurai, Erin Bradley, Iryna Bohovych, Bo Yarabe, Annemieke De Jong, Huib Ovaa, You Zhou, Oleh Khalimonchuk, Betzabet Quintanilla-Vega, Rodrigo Franco

Department of Biochemistry: Faculty Publications

Intracytoplasmic inclusions of protein aggregates in dopaminergic cells (Lewy bodies) are the pathological hallmark of Parkinson’s disease (PD). Ubiquitin (Ub), alpha [α]-synuclein, p62/sequestosome 1 and oxidized proteins are major components of Lewy bodies. However, the mechanisms involved in the impairment of misfolded/oxidized protein degradation pathways in PD are still unclear. PD is linked to mitochondrial dysfunction and environmental pesticide exposure. In this work, we evaluated the effect of the pesticide paraquat (PQ) and the mitochondrial toxin 1-methyl-4-phenylpyridinium (MPP⁺) on Ub-dependent protein degradation pathways. No increase in the accumulation of Ub-bound proteins or aggregates was observed in dopaminergic cells …

Potassium And The K+/H+ Exchanger Kha1p Promote Binding Of Copper To Apofet3p Multi-Copper Ferroxidase, Xiaobin Wu, Heejeong Kim, Javier Seravalli, Joseph J. Barycki, P. John Hart, David W. Gohara, Enrico Di Cera, Won Hee Jung, Daniel J. Kosman, Jaekwon Lee

Department of Biochemistry: Faculty Publications

Acquisition and distribution of metal ions support a number of biological processes. Here we show that respiratory growth of and iron acquisition by the yeast Saccharomyces cerevisiae relies on potassium (K⁺) compartmentalization to the trans-Golgi network via Kha1p, a K⁺/H⁺ exchanger. K⁺ in the trans-Golgi network facilitates binding of copper to the Fet3p multi-copper ferroxidase. The effect of K⁺ is not dependent on stable binding with Fet3p or alteration of the characteristics of the secretory pathway. The data suggest that K⁺ acts as a chemical factor in Fet3p maturation, a …

Analysis Of Oligomerization Properties Of Heme A Synthase Provides Insights Into Its Function In Eukaryotes, Samantha Swenson, Andrew Cannon, Nicholas J. Harris, Nicholas G. Taylor, Jennifer L. Fox, Oleh Khalimonchuk

Department of Biochemistry: Faculty Publications

Heme a is an essential cofactor for function of cytochrome c oxidase in the mitochondrial electron transport chain. Several evolutionarily conserved enzymes have been implicated in the biosynthesis of heme a, including the heme a synthase Cox15. However, the structure of Cox15 is unknown, its enzymatic mechanism and the role of active site residues remain debated, and recent discoveries suggest additional chaperone-like roles for this enzyme. Here, we investigated Cox15 in the model eukaryote Saccharomyces cerevisiae via several approaches to examine its oligomeric states and determine the effects of active site and human pathogenic mutations. Our results indicate that Cox15 …

Global Reach, Local Markets: The Challenges Of Leading Global Innovation, Sarah Higgins

Honors Theses

This Global Studies honors thesis addresses how managers and leaders of global firms manage innovation across multiple markets. Current research on multinational corporations provides an understanding of different kinds of innovation and the ways to attend to multiple markets. However, there is less documentation of how these innovation strategies are actually implemented on the ground and the tensions that these efforts might produce. Therefore, my research focuses in particular on the challenges and tensions faced by leaders of global firms as they implement transnational innovation strategies. This study is based upon in-depth interviews with 20 participants who held positions in …

การผลิตโปรตีเอสจากวัสดุเหลือทิ้งทางการเกษตรโดย Bacillus Pumilus, พัชราภรณ์ การงาน

Chulalongkorn University Theses and Dissertations (Chula ETD)

โปรตีเอสเป็นเอนไซม์ที่เร่งปฏิกิริยาการย่อยสลายโปรตีนให้เป็นกรดอะมิโน และเปปไทด์สายสั้นๆ ซึ่งสามารถนำไปประยุกต์ใช้ในการบำบัดน้ำเสียที่มีโปรตีนนมสูงจากร้านกาแฟได้ ซึ่งแบคทีเรียในสกุล Bacillus เป็นแบคทีเรียที่ผลิตโปรติเอสได้ดี โดย Bacillus pumilus มีความสามารถในการผลิตโปรติเอสปริมาณสูง ซึ่งคัดแยกได้จากบ่อน้ำพุร้อนทางภาคเหนือของประเทศไทย วิทยานิพนธ์นี้สนใจการผลิตโปรตีเอสจาก B. pumilus และศึกษาปัจจัยของแหล่งคาร์บอน และไนโตรเจน รวมถึงภาวะที่เหมาะสมต่อการผลิตโปรติเอส การผลิตโปรติเอสจาก B. pumilus มีค่ากิจกรรมของเอนไซม์โปรติเอสสูงที่สุด เท่ากับ 2.545 ยูนิตต่อมิลลิลิตร โดยเลี้ยงในอาหารสำหรับผลิตโปรติเอสประกอบด้วย 1 เปอร์เซ็นต์ โมลาส 1 เปอร์เซ็นต์ รำข้าว ภาวะในการเลี้ยงที่เหมาะสม คือ ค่าความเป็นกรด-ด่างเริ่มต้นในการเลี้ยงเท่ากับ 8.0 บ่มที่อุณหภูมิ 30 องศาเซลเซียส ความเร็วรอบ 250 รอบต่อนาที เป็นเวลา 24 ชั่วโมง ราคาอาหารเป็นหนึ่งในปัจจัยสำคัญสำหรับการผลิตโปรติเอส ดังนั้นการใช้วัสดุเหลือทิ้งทางการเกษตรเป็นแหล่งอาหารเลี้ยงเชื้อ เป็นการลดต้นทุนในการผลิตโปรตีเอส จึงเหมาะกับการนำไปประยุกต์ใช้ในระดับอุตสาหกรรม และการประยุกต์ใช้ในการบำบัดน้ำเสีย

Alcoholic Vs Non-Alcoholic Fatty Liver In Rats: Distinct Differences In Endocytosis And Vesicle Trafficking Despite Similar Pathology, Karuna Rasineni, Daniel D. Penrice, Sathish Kumar Natarajan, Mark A. Mcniven, Benita L. Mcvicker, Kusum K. Kharbanda, Carol A. Casey, Edward N. Harris

Department of Biochemistry: Faculty Publications

Background: Non-alcoholic and alcoholic fatty liver disease (NAFLD and AFLD, respectively) are major health problems, as patients with either condition can progress to hepatitis, fibrosis, and cirrhosis. Although histologically similar, key differences likely exist in these two models. For example, altered content of several vesicle trafficking proteins have been identified in AFLD, but their content in NAFLD is unknown. In this study, we compared select parameters in NAFLD and AFLD in a rat model.

Methods: We fed either Lieber- DeCarli liquid control or alcohol-containing (35 % as calories) diet (AFLD model) or lean or high-fat (12 or 60 % derived …

Atomic Resolution Experimental Phase Information Reveals Extensive Disorder And Bound 2-Methyl-2,4-Pentanediol In Ca2+-Calmodulin, Jiusheng Lin, Henry Van Den Bedem, Axel T. Brunger, Mark A. Wilson

Department of Biochemistry: Faculty Publications

Calmodulin (CaM) is the primary calcium signaling protein in eukaryotes and has been extensively studied using various biophysical techniques. Prior crystal structures have noted the presence of ambiguous electron density in both hydrophobic binding pockets of Ca²⁺-CaM, but no assignment of these features has been made. In addition, Ca²⁺-CaM samples many conformational substates in the crystal and accurately modeling the full range of this functionally important disorder is challenging. In order to characterize these features in a minimally biased manner, a 1.0 A resolution single-wavelength anomalous diffraction data set was measured for selenomethionine-substituted Ca²⁺-CaM. …

An Evaluation Of New And Established Methods To Determine T-Dna Copy Number And Homozygosity In Transgenic Plants, Katarzyna Glowacka, Johannes Kromdijk, Lauriebeth Leonelli, Krishna K. Niyogi, Tom E. Clemente, Stephen P. Long

Department of Biochemistry: Faculty Publications

Stable transformation of plants is a powerful tool for hypothesis testing.Arapid and reliable evaluation method of the transgenic allele for copy number and homozygosity is vital in analysing these transformations.Here the suitability of Southern blot analysis, thermal asymmetric interlaced (TAIL-)PCR, quantitative (q)PCR and digital droplet (dd)PCR to estimate T-DNA copy number, locus complexity and homozygosity were compared in transgenic tobacco. Southern blot analysis and ddPCR on three generations of transgenic offspring with contrasting zygosity and copy number were entirely consistent, whereas TAIL-PCR often underestimated copy number. qPCR deviated considerably from the Southern blot results and had lower precision and higher …

Encapsulation Of Natural Colorant In Rice Flour Granules, Pornpan Natthawut

Chulalongkorn University Theses and Dissertations (Chula ETD)

The trend of using natural colorants replaced synthetic colorants was increasing in food and cosmetic application. However, the natural colorants had disadvantage on color instability. Usually the color of anthocyanin changes quite quickly after the materials are exposed to various environmental factors including moisture, air and light. In this work, rice granules were used to stabilize roselle, butterfly pea and curcumin extracts. The stabilized natural colorants were then subjected to stability evaluation at various temperatures (room temperature, 4oC and 40oC) for up to 1000 h, and also under light-exposed and light-proof conditions for up to 100 h. The monitoring of …

Chlamydia Trachomatis Transformants Show A Significant Reduction In Rates Of Invasion Upon Removal Of Key Tarp Domains, Christopher Parrett

Electronic Theses and Dissertations

Chlamydia trachomatis is an obligate, intracellular bacterium which is known to cause multiple human infections including nongonococcal urethritis (serovars D-K), lymphogranuloma venereum (serovars L1, L2, L3) and trachoma (serovars A-C). The infectious form of the bacterium, called the elementary body (EB), harbors a type III secreted effector known as Tarp (translocated actin recruiting phosphoprotein) which is a candidate virulence factor and is hypothesized to play a role in C. trachomatis' ability to invade and grow within epithelial cells in a human host. C. trachomatis L2 Tarp harbors five unique protein domains which include the Phosphorylation Domain, the Proline Rich Domain, …