Biotechnology Commons^™

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 …

Insights Into Triterpene Metabolism In Model Monocotyledonous And Oilseed Plants Genetically Engineered With Genes From Botryococcus Braunii, Chase F. Kempinski

Theses and Dissertations--Plant and Soil Sciences

Isoprenoids are one of the most diverse classes of natural products and are all derived from universal five carbon, prenyl precursors. Squalene and botryococcene are linear, hydrocarbon triterpenes (thirty carbon compounds with six prenyl units) that have industrial and medicinal values. Squalene is produced by all eukaryotes as it is the first committed precursor to sterols, while botryococcene is uniquely produced by the green algae, Botryococcus braunii (race B). Natural sources for these compounds exist, but there is a desire for more renewable production platforms. The model plant Arabidopsis thaliana was engineered to accumulate botryococcene and squalene in its oil …

Probing The Plant Cell Wall With Herbicides: A Chemical Genetics Approach, Chad B. Brabham

Theses and Dissertations--Plant and Soil Sciences

The primary cell wall is a highly organized multi-layered matrix of polysaccharides (cellulose, hemi-cellulose, and pectin). The ability of the rigid cell wall to sufficiently loosen to allow growth is a complex process that differs considerably between grasses monocots and dicots. Cellulose is the major structural component required for anisotropic cell expansion and is synthesized by CELLULOSE SYNTHASE A (CesA) proteins. Here, our objectives were two-fold: 1) dissect cell walls and cellulose biosynthesis in dicots and grasses using chemical biology and reverse genetic approaches 2) characterize and classify the inhibitory mechanisms of cellulose biosynthesis inhibitors (CBIs). A reverse genetics TILLING …

A Novel Transcription Factor In Arabidopsis Thaliana Abiotic Stress Response, Achira S. Weerathunga Arachchilage

LSU New Orleans Theses and Dissertations

Plants respond to environmental stress by altering their gene expression. Under stress conditions some genes are activated and some genes are repressed. Even though a lot of work has been done to understand mechanisms of gene activation under abiotic stress very little information is available on how stress responsive genes are kept repressed under normal growth conditions. Recent work has revealed that plants use transcriptional repression as common mechanism of gene repression. Transcriptional repression is achieved by recruitment co-repressor complexes to the target genes. Recent studies have revealed that the co-repressor LUH complexes with SLK1 and SLK2 to silence Arabidopsis …

Production And Harvest Of Microalgae In Wastewater Raceways With Resource Recycling, Alexander Colin Roberts

Master's Theses

Microalgae can be grown on municipal wastewater media to both treat the wastewater and produce feedstock for algae biofuel production. However the reliability of treatment must be demonstrated, as well as high areal algae productivity on recycled wastewater media and efficient sedimentation harvesting. This processes was studied at pilot scale in the present research.

A pilot facility was operated with nine CO₂-supplemented raceway ponds, each with a 33-m² surface area and a 0.3-m depth, continuously from March 6, 2013 through September 24, 2014. The ponds were operated as three sets of triplicates with two sets continuously fed …

Pertumbuhan Dan Perkembangan Tunas Nilam Var. Lhoukseumawe Dari Jenis Eksplan Dengan Sitokinin Yang Berbeda Secara In Vitro, Erni Suminar, Intan Ratna Dewi Anjarsari, Anne Nuraini, Hapizhah

Jurnal Kultivasi

Tujuan dari penelitian ini adalah untuk mendapatan metode perbanyakan tanaman nilam melalui teknik in vitro dengan meng-gunakan jenis eksplan serta jenis dan konsentrasi sitokinin yang berbeda. Percobaan dilaksanakan di Laboratorium Teknologi Benih Fakultas Pertanian Universitas Padjadjaran dari bulan Maret sampai dengan Juni 2012. Rancangan percobaan menggunakan Rancangan Acak Lengkap pola faktorial yang terdiri dua faktor dan tiga kali ulangan. Faktor pertama adalah tipe eksplan (mata tunas, pucuk, dan daun). Faktor kedua adalah tipe dan konsentrasi sitokinin (0 mg L-1, 0,5 mgL-1 BAP, 1,0 mgL-1 BAP0,5 mgL-1 zeatin, and 1,0 mgL-1 zeatin). Hasil percobaan menunjukkan bahwa eksplan pucuk dengan penggunaan 0.5 …

Anaerobic Detoxification Of Acetic Acid In A Thermophilic Ethanologen, A Joe Shaw, Bethany B. Miller, Stephen R. Rogers, William Robert Kenealy, Alex Meola, Ashwini Bhandiwad, W Ryan Sillers, Indraneel Shikhare, David Hogsett, Christopher Herring

Dartmouth Scholarship

The liberation of acetate from hemicellulose negatively impacts fermentations of cellulosic biomass, limiting the concentrations of substrate that can be effectively processed. Solvent-producing bacteria have the capacity to convert acetate to the less toxic product acetone, but to the best of our knowledge, this trait has not been transferred to an organism that produces ethanol at high yield. We have engineered a five-step metabolic pathway to convert acetic acid to acetone in the thermophilic anaerobe Thermoanaerobacterium saccharolyticum.

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Reducing The Negative Human-Health Impacts Of Bioenergy Crop Emissions Through Region-Specific Crop Selection, William Christian Porter, Todd N. Rosenstiel, Alex Guenther, Jean-Francois Lamarque, Kelley Barsanti

Biology Faculty Publications and Presentations

An expected global increase in bioenergy-crop cultivation as an alternative to fossil fuels will have consequences on both global climate and local air quality through changes in biogenic emissions of volatile organic compounds (VOCs). While greenhouse gas emissions may be reduced through the substitution of next-generation bioenergy crops such as eucalyptus, giant reed, and switchgrass for fossil fuels, the choice of species has important ramifications for human health, potentially reducing the benefits of conversion due to increases in ozone (O3) and fine particulate matter (PM2.5) levels as a result of large changes in biogenic emissions. Using the Community Earth System …

Assessment Of Variation In Susceptibility Of The Fall Armyworm, Spodoptera Frugiperda (J.E. Smith) (Lepidoptera: Noctuidae), To Bacillus Thuringiensis Toxins, Karen Ferreira Da Silva

Department of Entomology: Dissertations, Theses, and Student Research

The fall armyworm, Spodoptera frugiperda, is a polyphagous insect pest affecting multiple crops. Fall armyworm is managed with insecticides and corn hybrids expressing insecticidal proteins derived from Bacillus thuringiensis. The early detection of insect resistance is important for making appropriate management decisions informs IPM and IRM recommendations.

The objective of the first study was to establish baseline susceptibility of fall armyworm populations to the Cry1F Bt insecticidal protein, emphasizing collections from locations where fall armyworm overwinters in the U.S. Fall armyworm neonates were exposed to artificial diet treated with increasing Cry1F concentrations, and mortality and growth inhibition were …

Increasing Expression Of Hepatitis B Surface Antigen In Maize Through Breeding, Erin Suzanne Miller

Master's Theses

The hepatitis B virus (HBV) is a common virus, with two billion people infected worldwide. It causes approximately 600,000 deaths each year, despite the availability of an effective vaccine since 1982. Maize as a platform for oral vaccination can supply a heat stable vaccine, which does not require syringes or trained personnel to administer. The Hepatitis B Surface antigen was transformed into maize and this seed was used to evaluate expression levels through the breeding process. The transgene was transferred into two elite maize inbreds by backcrossing. Highest expressing ears were selected each generation until approximately 99% commercial parent was …

Target-Directed Biosynthetic Evolution: Redirecting Plant Evolution To Genomically Optimize A Plant’S Pharmacological Profile, Dustin Paul Brown

Theses and Dissertations--Neuroscience

The dissertation describes a novel method for plant drug discovery based on mutation and selection of plant cells. Despite the industry focus on chemical synthesis, plants remain a source of potent and complex bioactive metabolites. Many of these have evolved as defensive compounds targeted on key proteins in the CNS of herbivorous insects, for example the insect dopamine transporter (DAT). Because of homology with the human DAT protein some of these metabolites have high abuse potential, but others may be valuable in treating drug dependence. This dissertation redirects the evolution of a native Lobelia species toward metabolites with greater activity …

The Exometabolome Of Clostridium Thermocellum Reveals Overflow Metabolism At High Cellulose Loading, Evert K. Holwerda, Philip G. Thorne, Daniel G. Olson, Daniel Amador-Noguez, Nancy L. Engle, Timothy J. Tschaplinski, Johannes P. Van Dijken, Lee R. Lynd

Dartmouth Scholarship

BackgroundClostridium thermocellum is a model thermophilic organism for the production of biofuels from lignocellulosic substrates. The majority of publications studying the physiology of this organism use substrate concentrations of ≤10 g/L. However, industrially relevant concentrations of substrate start at 100 g/L carbohydrate, which corresponds to approximately 150 g/L solids. To gain insight into the physiology of fermentation of high substrate concentrations, we studied the growth on, and utilization of high concentrations of crystalline cellulose varying from 50 to 100 g/L by C. thermocellum. .

Overexpression Of Patatin-Related Phospholipase Aiiiβ Altered The Content And Composition Of Sphingolipids In Arabidopsis, Maoyin Li, Jennifer E. Markham, Xuemin Wang

Department of Biochemistry: Faculty Publications

In plants, fatty acids are primarily synthesized in plastids and then transported to the endoplasmic reticulum (ER) for synthesis of most of the complex membrane lipids, including glycerolipids and sphingolipids. The first step of sphingolipid synthesis, which uses a fatty acid and a serine as substrates, is critical for sphingolipid homeostasis; its disruption leads to an altered plant growth. Phospholipase As have been implicated in the trafficking of fatty acids from plastids to the ER. Previously, we found that overexpression of a patatin-related phospholipase, pPLAIIIβ, resulted in a smaller plant size and altered anisotropic cell expansion. Here, we determined the …

Genome Wide Association Mapping Of Grain Arsenic, Copper, Molybdenum And Zinc In Rice (Oryza Sativa L.) Grown At Four International Field Sites, Gareth J. Norton, Alex Douglas, Brett Lahner, Elena Yakubova, Mary Lou Guerinot, Shannon R.M Pinson, Lee Tarpley, George C. Eizenga, Steve P. Mcgrath, Fang-Jie Zhao

Dartmouth Scholarship

The mineral concentrations in cereals are important for human health, especially for individuals who consume a cereal subsistence diet. A number of elements, such as zinc, are required within the diet, while some elements are toxic to humans, for example arsenic. In this study we carry out genome-wide association (GWA) mapping of grain concentrations of arsenic, copper, molybdenum and zinc in brown rice using an established rice diversity panel of ~300 accessions and 36.9 k single nucleotide polymorphisms (SNPs). The study was performed across five environments: one field site in Bangladesh, one in China and two in the US, with …

Arabidopsis Accelerated Cell Death 11, Acd11, Is A Ceramide-1-Phosphate Transfer Protein And Intermediary Regulator Of Phytoceramide Levels, Dhirendra K. Simanshu, Xiuhong Zhai, David Munch, Daniel Hofius, Jennifer E. Markham, Jacek Bielawski, Alicja Bielawska, Lucy Malinina, Julian G. Molotkovsky, John W. Mundy, Dinshaw J. Patel, Rhoderick E. Brown

Department of Biochemistry: Faculty Publications

The accelerated cell death 11 (acd11) mutant of Arabidopsis provides a genetic model for studying immune response activation and localized cellular suicide that halt pathogen spread during infection in plants. Here, we elucidate ACD11 structure and function and show that acd11 disruption dramatically alters the in vivo balance of sphingolipid mediators that regulate eukaryotic-programmed cell death. In acd11 mutants, normally low ceramide-1- phosphate (C1P) levels become elevated, but the relatively abundant cell death inducer phytoceramide rises acutely. ACD11 exhibits selective intermembrane transfer of C1P and phyto-C1P. Crystal structures establish C1P binding via a surface-localized, phosphate headgroup recognition center …

Estimating The Frequency Of Cry1f Resistance In Field Populations Of The European Corn Borer (Lepidoptera: Crambidae), Blair D. Siegfried, Murugesan Rangasamy, Haichuan Wang, Terence A. Spencer, Chirakkal V. Haridas, Brigitte Tenhumberg, Douglas V. Sumerford, Nicholas P. Storer

School of Biological Sciences: Faculty Publications

Background: Transgenic corn hybrids that express toxins from Bacillus thuringiensis (Bt) have suppressed European corn borer populations and reduced the pest status of this insect throughout much of the US corn belt. A major assumption of the high-dose/refuge strategy proposed for insect resistance management and Bt corn is that the frequency of resistance alleles is low so that resistant pests surviving exposure to Bt corn will be rare. Results: The frequency of resistance to the Cry1F Bt toxin was estimated using two different screening tools and compared with annual susceptibility monitoring based on diagnostic bioassays and LC_{50 …}

Tracking The Cellulolytic Activity Of Clostridium Thermocellum Biofilms, Alexandru Dumitrache, Gideon M. Wolfaardt, David Allen, Steven N. Liss, Lee R. Lynd

Dartmouth Scholarship

Microbial cellulose conversion by Clostridium thermocellum 27405 occurs predominantly through the activity of substrate-adherent bacteria organized in thin, primarily single cell-layered biofilms. The importance of cellulosic surface exposure to microbial hydrolysis has received little attention despite its implied impact on conversion kinetics.

Development Of A Metabolomic Method To Define The Phenylalanome In Arabidopsis Thaliana, Cole G. Wunderlich, Clint Chapple, Xu Li

The Summer Undergraduate Research Fellowship (SURF) Symposium

In the study of metabolomics, one of the greatest challenges can be accurately identifying compounds detected in biological extracts, especially when standards are not readily available. Current metabolomic methods are also limited in that they provide little to no information about a compound’s metabolic origin. In this study, we sought to address these issues by developing a novel metabolomic method that employs stable isotope feeding, LC-MS, Xcms, and an analytical software algorithm to study the ‘phenylalanome’ of Arabidopsis thaliana. Using this approach we were able to develop a method that, based on current results, is capable of detecting over …

Pembentukan Ubi Mikro Kentang Kultivar Granola Dengan Penggunaan Jenis Dan Konsentrasi Zat Inhibitor, Rifky Yusuf, Erni Suminar

Jurnal Kultivasi

The aim of the experiment was to obtain the concentration of inhibitors such as coumarin and paclobutrazol on the growth of micro-cuttings and in vitro tuberization of potato micro tuber. It can be used as source of high quality seeds.Experiments was conducted in the Tissue Culture laboratory Faculty of Agriculture, University of Padjadjaran in Jatinangor, Sumedang, from March 2011 to July 2011. Randomized Complete Design (RCD) was used in this experiment. They areconsist of 12 treatments with three replications, with treatment as follows: coumarin (30 mg L-1, 45 mg L-1, 60 mg L-1, 75 mg L-1, 90 mg L-1, 105 …

A Morphological Analysis Of A Hybrid Swarm Of Native Ulmus Rubra And Introduced U. Pumila (Ulmaceae) In Southeastern Nebraska, Christian Elowsky, Ingrid E. Jordon-Thaden, Robert B. Kaul

Nebraska Center for Biotechnology: Faculty and Staff Publications

The parental species and hybrid swarm of native Ulmus rubra Muhl. and the introduced, naturalized and weedy U. pumila L. were investigated in a 65-km transect in Lancaster, Saunders, and Butler counties in Nebraska. Thirty-two trees of U. rubra, 32 of U. pumila, and 50 of the hybrid swarm were sampled for leaves and buds and subsampled for flowers and fruits. Leaves were measured for petiole length, blade length, width, primary and secondary teeth per cm, number of secondary teeth per primary tooth, and texture. Buds were scored for color and distribution of trichomes. Flowers were sampled for stamen counts …

Overexpression/Silencing Of Selected Soybean Genes Alters Resistance To Pathogens, Mohamed H. El-Habbak

Theses and Dissertations--Plant Pathology

Plant diseases remain a major obstruction to meeting the world’s increased demand for soybean oil and protein. Reducing the losses caused by diseases in order to improve crop production is a high priority for agricultural research. The need for novel strategies for plant disease control cannot be overstated. In the present study, selected defense-related genes were silenced and/or overexpressed in soybean using a virus-based vector and the resultant plants were tested for their responses to pathogens. The first part of the study focused on Rps1k (Resistance to Phytophthora sojae) gene. The two conserved domains encoding ‘P-Loop NTPase’ and ‘PLN03210’ …

Carbon Farming In Wa, Department Of Agriculture And Food, Western Australia, Department Of Regional Development And Lands, Western Australia

Natural resources published reports

Fact sheets:

1. Applying biochar to improve soil quality and crop productivity

2. Reducing methane emissions from cattle using feed additives

3. Cattle breeding for lower greenhouse gas emissions

4. Managing genetic codes to reduce methane emissions from cattle

5. Managing pastures and stocking rate to reduce methane emissions from cattle

6. Claying as a method of increasing soil carbon content

7. Green and brown manuring as part of carbon farming

8. Permanent environmental plantings to earn carbon credits

9. Nitrous oxide emissions from agricultural soils

10. Sheep genetics in methane reduction

11. Managing sheep pastures to reduce methane production …

Connections Between Sphingosine Kinase And Phospholipase D In The Abscisic Acid Signaling Pathway In Arabidopsis, Liang Guo, Girish Mishra, Jennifer E. Markham, Maoyin Li, Amanda Tawfall, Ruth Welti, Xuemin Wang

Department of Biochemistry: Faculty Publications

Background: Sphingosine kinase (SPHK) and phospholipaseD(PLD) produce different lipid mediators involved in abscisic acid (ABA) response.

Results: Ablation of SPHKs and PLDα1 attenuates ABA-induced production of LCBPs and PA. Phyto-S1P closes stomata in sphk1, sphk2, but not in pldα1, whereas PA closes stomata in all mutants.

Conclusion: SPHK acts upstream of PLDα1, whereas PLDα1 promotes SPHK.

Significance: The roles of lipid messengers in the ABA signaling pathway are clarified.

Modeling An Improvement In Phosphorus Utilization In Tropical Agriculture, David Edelstein, David J. Tonjes

Technology & Society Faculty Publications

Studies of Terra Preta soils have generated interest in recreating their fertility elsewhere. Much of the research has focused on soil amendment charcoal (“biochar”). Terra Preta also contains bone fragments, producing a high concentration of phosphorus. Some forecast worldwide declines in phosphorus supplies, and better agricultural system management is required to improve phosphorus use efficiency. A conceptual model is offered to consider the influence of charcoal on bioavailability of phosphorus. The model describes a system where improvements in the chemical and biological condition of the soil result in increased phosphorus availability and cycling. Mechanisms of phosphorus/charcoal interaction are considered, and …

Propagation System Manual, Margaret Kane

Hydroponic Center

Contained in this manual you will find the required materials, assembly instructions, operation procedures and safety protocols to construct, run and maintain a hydroponic propagation system.

The Propagation System is designed to provide an environment for seeds to germinate with optimal conditions for seedling growth. The purpose of the Propagator is to create an environment that is suitable for seedlings to develop a strong root system and uprooted stem so they can be transplanted into a new system where they are able to fully develop and be harvested. This procedure introduces plants, vegetables, and fruits to a new system of …

A Novel Gene Silencing Vector For Plant Genomics And Biotechnology, Dustin Hoover, Scott Nicholson, Vibha Srivastava

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

Gene silencing is a process of suppressing activity of specific genes by producing “interfering” RNA encoded by foreign genes. This process serves as the principle of genetic modification in plants and animals, which is an important tool in genomics and biotechnology, allowing scientists to manipulate organisms to better meet human demands. New approaches of gene silencing may enable improvements on current practices of genetic modification, and broaden the application and impact of gene silencing in biotechnology. Recently, a novel vector design consisting of the transcription of short gene fragments lacking transcription termination signals was demonstrated to be effective in partial …

Purification And Analysis Of Bacterial-And Plant-Derived Scfv Antibodies For The Detection Of Cucumber Mosaic Virus, Wong C.F.Y.

Student Works (2010-2019)

abstract N/A

Biolistic Transformation Of Oil Palm Using The Phosphomannose Isomerase (Pmi) Gene As The Positive Selectable Marker., Bahariah Bohari

Student Works (2010-2019)

The selectable marker system based on the Escherichia coli phosphomannose isomerase (pmi) gene was adapted for genetic transformation of oil palm. This system makes use of the pmi gene that encodes phosphomannose isomerase, which converts mannose-6-phosphate to fructose-6-phosphate and uses mannose as the selection agent. This is to anticipate the future requirement of using non-antibiotic resistance genes for the commercialization of transgenic oil palm. The use of antibiotic or herbicide-based selection systems has caused much public concern due to inadequate knowledge of the agents’ impact on the environment and on human health. In this study, four transformation vectors, namely pMI3, …

Studies On The Bioactive Properties Of Selected Plant And Fermented Extracts., Saravana Kumar Sinniah

Student Works (2010-2019)

The aim of this study was to isolate protein/peptide and secondary metabolites from plant and fermented sources. The extracts were used to test for antimicrobial and antioxidant properties. In the current study protein/peptide extracts of Allium sativum showed promising results with a 15mm inhibition zone against Escherichia coli, 28mm against Staphylococcus aureus, 16.3mm against Bacillus cereus and 9mm against Pseudomonas aeruginosa. The minimum bacteriocidal concentration (MBC) results showed that Allium sativum protein/peptide extract is only bacteriostatic and not bacteriocidal. Andrographis paniculata showed the most promising result for the ethanolic extracts with an inhibition zone of 14mm against B. cereus, 11.5mm …

Agrobacterium-Mediated Transformation Of Cryptocoryne Willisii With Gus And Mgfp 5 Reporter Genes / Wong Sher Ming, Wong Sher Ming

Student Works (2010-2019)

In this study, a simple method using Murashige and Skoog basal medium (1962) supplemented with 6-BA alone was developed for direct multiple shoots induction from shoot base culture of C. willisii. Shoots proliferation and elongation were achieved on a single medium without the need of subculturing. The optimum concentration of 6-BA for direct shoots regeneration of C. willisii was investigated. It was found to be 6 mg/ L. Optimum media was then used for regenerate and propagate of transformed C. willisii. On the other hand, ransformation of C. willisii with Agrobacterium tumefaciens strain LBA4404 harbouring pCAMBIA1304 was performed. Effects on …