Biotechnology Commons^™

Drosophila As An Emerging Model To Study Metastasis, Madhuri Kango-Singh, Georg Halder

Biology Faculty Publications

Metastasis is the primary cause of human cancer-related deaths. Two recent studies describe a system for testing how multiple genetic events synergize to promote neoplastic growth and metastasis in Drosophila, paving the way for systematic approaches to understanding metastasis using the powerful tools of Drosophila genetics.

Morphological Characteristics And Species Separation Of Hawaiian Postlarval Amphidromous Fishes, M. Eric Benbow, Albert J. Burky, Carl M. Way

Biology Faculty Publications

There are five Hawaiian amphidromous fishes (Gobiidae: Lentipes concolor, Awaous guamensis, Sicyopterus stimpsoni, Stenogobius hawaiiensis; Eleotridae: Eleotris sandwicensis). Amphidromous adults deposit eggs on the stream bottom. After hatching, larvae drift to the ocean for growth followed by postlarval migration back into the streams. Postlarvae were collected to construct a dichotomous identification key based on 12 morphological measures and ray counts from four fins. Overall, the presense of fused pelvic fins, standard length (SL), and fin ray numbers were the most useful in species separation. Gobies were separated from the eleotrid by the former having fused …

Computational Protein Biomarker Prediction: A Case Study For Prostate Cancer, Michael Wagner, Dayanand N. Naik, Alex Pothen, Srinivas Kasukurti, Raghu Ram Devineni, Bao-Ling Adam, O. John Semmes, George L. Wright Jr.

Mathematics & Statistics Faculty Publications

Background: Recent technological advances in mass spectrometry pose challenges in computational mathematics and statistics to process the mass spectral data into predictive models with clinical and biological significance. We discuss several classification-based approaches to finding protein biomarker candidates using protein profiles obtained via mass spectrometry, and we assess their statistical significance. Our overall goal is to implicate peaks that have a high likelihood of being biologically linked to a given disease state, and thus to narrow the search for biomarker candidates.

Results: Thorough cross-validation studies and randomization tests are performed on a prostate cancer dataset with over 300 patients, obtained …

Initial State Of The Drosophila Eye Before Dorsoventral Specification Is Equivalent To Ventral, Amit Singh, Kwang-Wook Choi

Biology Faculty Publications

Dorsoventral (DV) patterning is crucial for eye development in invertebrates and higher animals. DV lineage restriction is the primary event in undifferentiated early eye primordia of Drosophila. InDrosophila eye disc, a dorsal-specific GATA family transcription factor pannier (pnr) controls Iroquois-Complex (Iro-C) genes to establish the dorsal eye fate whereas Lobe (L), which is involved in controlling a Notch ligand Serrate (Ser), is specifically required for ventral growth. However, fate of eye disc cells before the onset of dorsal expression of pnr and Iro-C is not known. We show that L/Ser …

Life Cycle Of A Torrenticolous Hawaiian Chironomid (Telmatogeton Torrenticola): Stream Flow And Microhabitat Effects, M. Eric Benbow, Albert J. Burky, Carl M. Way

Biology Faculty Publications

In this study we documented the instar densities and life cycle of Telmatogeton torrenticola Terry (Chironomidae : Telmatogetoninae) from Kinihapai Stream, Maui, Hawaii. Greatest larval densities of this midge are found on substrates of high velocity, shallow flows of cascades, and splash zones of waterfalls, with lower densities in riffles. In the summer of 1994 we compared the effects of two microhabitats (termed optimal and suboptimal) on inter-instar density and relative abundance. In a second year (1995), we evaluated the effect of long-term reduced stream flow on these variables only in optimal microhabitats. A significant reduction in stream flow from …

Comparison Of Methods For Dna Isolation From Food Samples For Detection Of Shiga Toxin-Producing Escherichia Coli By Real-Time Pcr, Loree C. Heller, Carisa R. Davis, K. Kealy Peak, David Wingfield, Andrew C. Cannons, Philip T. Amuso, Jacqueline Cattani

Bioelectrics Publications

In this study, food samples were intentionally contaminated with Escherichia coli O157:H7, and then DNA was isolated by using four commercial kits. The isolated DNA samples were compared by using real-time PCR detection of the Shiga toxin genes. The four kits tested worked similarly.

Shar-Pei Mediates Cell Proliferation Arrest During Imaginal Disc Growth In Drosophila, Madhuri Kango-Singh, Riitta Nolo, Chunyao Tao, Patrik Verstreken, P. Robin Hiesinger, Hugo J. Bellen, Georg Halder

Biology Faculty Publications

During animal development, organ size is determined primarily by the amount of cell proliferation, which must be tightly regulated to ensure the generation of properly proportioned organs. However, little is known about the molecular pathways that direct cells to stop proliferating when an organ has attained its proper size. We have identified mutations in a novel gene, shar-pei, that is required for proper termination of cell proliferation during Drosophila imaginal disc development. Clones of shar-pei mutant cells in imaginal discs produce enlarged tissues containing more cells of normal size. We show that this phenotype is the result of both …

Electrically Mediated Delivery Of Vector Plasmid Dna Elicits An Antitumor Effect, L. Heller, D. Coppola

Bioelectrics Publications

In vivo electroporation is an efficient means of increasing plasmid DNA delivery to normal tissues, such as skin and muscle, as well as directly to tumors. In the experiments described here, plasmid DNA was delivered by in vivo electroporation to B16 mouse melanomas using two very different pulsing protocols. Reporter expression increased 21- or 42-fold, respectively with electroporation over injection alone. The growth of experimental melanomas with an approximate diameter of 4 mm on the day of treatment was monitored after electroporation delivery of reporter plasmid DNA. Remarkably, short-term complete regressions using one of these pulsing protocols occurred in up …

Eye Suppression, A Novel Function Of Teashirt, Requires Wingless Signaling, Amit Singh, Madhuri Kango-Singh, Y. Henry Sun

Biology Faculty Publications

Teashirt (tsh) encodes a Drosophila zinc-finger protein. Misexpression of tsh has been shown to induce ectopic eye formation in the antenna. We report that tsh can suppress eye development. This novel function of tsh is due to the induction of homothorax (hth), a known repressor of eye development, and requires Wingless (WG) signaling. Interestingly, tsh has different functions in the dorsal and ventral eye, suppressing eye development close to the ventral margin, while promoting eye development near the dorsal margin. It affects both growth of eye disc and retinal cell differentiation.

Effect Of Electrically Mediated Intratumor And Intramuscular Delivery Of A Plasmid Encoding Ifn Α On Visible B16 Mouse Melanomas, Loree C. Heller, Stephanie F. Ingram, M. Lee Lucas, Richard A. Gilbert, Richard Heller

Bioelectrics Publications

Interferon α may be used as a single agent therapy for metastatic malignant melanoma or as an adjuvant to chemotherapy. Delivery of interferon α by gene therapy offers an alternative to recombinant protein therapy. Electrically mediated delivery enhances plasmid expression in a number of tissues, for instance skin, liver, muscle and tumors including melanomas. Here we compare the effect of delivery of a plasmid encoding mouse interferon α on growth of visible B16 mouse melanomas following electrically mediated delivery to muscle or directly to the tumor. Intratumoral delivery of interferon α plasmid not only slows melanoma growth, but induces complete, …

Il-12 Plasmid Delivery By In Vivo Electroporation For The Successful Treatment Of Established Subcutaneous B16.F10 Melanoma, M. Lee Lucus, Loree Heller, Domenico Coppola, Richard Heller

Bioelectrics Publications

Interleukin-12 (IL-12) has been used in numerous immunotherapy protocols against melanoma. However, delivery of IL-12 in the form of recombinant protein can result in severe toxicity, and gene therapy has had limited success against B16.F10 murine melanoma. The purpose of this study was to examine the effectiveness of in vivo electroporation for the delivery of plasmid DNA encoding IL-12 as an antitumor agent against B16.F10 melanoma. We treated mice bearing established B16.F10 melanoma tumors with intratumoral (i.t.) or intramuscular (i.m.) injections of a plasmid encoding IL-12, followed by in vivo electroporation. For i.t. treatments, we used an applicator containing six …

Functional Requirement Of Aquaporin-5 In Plasma Membranes Of Sweat Glands, Lene N. Nejsum, Tae-Hwan Kwon, Uffe B. Jensen, Ornella Fumagalli, Jørgen Frøkiaer, Carissa M. Krane, Anil G. Menon, Landon S. King, Peter C. Agre, Søren Nielsen

Biology Faculty Publications

The distribution and function of aquaporins (AQPs) have not previously been defined in sweat glands. In this study, AQP1, AQP3, and AQP5 mRNA were demonstrated in rat paw by reverse transcription (RT)–PCR, but AQP2 and AQP4 were not. AQP1, AQP3, and AQP5 protein were confirmed in these tissues by immunoblotting. AQP1 was identified in capillary endothelial cells by immunohistochemical labeling, but not in sweat glands or epidermis. Abundant AQP3 expression was seen in basal levels of epidermis, but not in sweat glands. AQP2 and AQP4 were not observed in either skin or sweat glands. Immunohistochemical labeling revealed abundant AQP5 in …

Genetic Variability Studies In A Selected Population Of Azadirachta Excelsa (Sentang) With Aflp Marker Technology, Chiew Foan Chin

Student Works (2000-2009)

A maternal half-s/b population of Azadirachta excelsa was established from seeds in a randomised complete block design with three replicates. 100 to 200 seeds were collected from each of the 13 selected mother trees. The mother trees were categorised into three groups i.e. large, medium and small according to their girth sizes. 50 progeny seedlings from each scedlot per replicate were planted in the experimental field. Genetic studies on phenotypic measurements and molecular marker analysis were carried out on the established A. excelsa population. Measurements of quantitative traits including diameter breast height (Dbh) and total height were made. Mean Dbh …

A Note On Cascade Climbing Of Migrating Goby And Shrimp Postlarvae In Two Maui Streams, M. Eric Benbow, Leslie Luchar Orzetti, Mollie D. Mcintosh, Albert J. Burky

Biology Faculty Publications

In this study, we documented cascade climbing rates of 133 and 230 postlarvae of Lentipes concolor (O‘opu alamo‘o) and Atyoida bisulcata (Opae kahaole), respectively, from two streams on the island of Maui, Hawaii. Climbing measurements and observations were made of postlarvae at the water-substrate interface in cascade habitats of constricted water flow. Both species were observed to move in short bursts of forward progression within or above the pulsing water-substrate interface. Goby postlarval climbing rates ranged from 0.04 – 1.50 cm s–1 and were slower than shrimp rates which ranged from 0.30 – 3.06 cm s–1. The high variability is …

Salivary Acinar Cells From Aquaporin 5-Deficient Mice Have Decreased Membrane Water Permeability And Altered Cell Volume Regulation, Carissa M. Krane, James E. Melvin, Ha-Van Nguyen, Linda Richardson, Jennifer E. Towne, Thomas Doetschman, Anil G. Menon

Biology Faculty Publications

Aquaporins (AQPs) are channel proteins that regulate the movement of water through the plasma membrane of secretory and absorptive cells in response to osmotic gradients. In the salivary gland, AQP5 is the major aquaporin expressed on the apical membrane of acinar cells. Previous studies have shown that the volume of saliva secreted by AQP5-deficient mice is decreased, indicating a role for AQP5 in saliva secretion; however, the mechanism by which AQP5 regulates water transport in salivary acinar cells remains to be determined. Here we show that the decreased salivary flow rate and increased tonicity of the saliva secreted byAqp5 …

Studies On The Formation Of Dna-Cationic Lipid Composite Films And Dna Hybridization In The Composites, Murali Sastry, Vidya Ramakrishnan, Mrunalini Pattarkine, Krishna N. Ganesh

Faculty Works

The formation of composite films of double-stranded DNA and cationic lipid molecules (octadecylamine, ODA) and the hybridization of complementary single-stranded DNA molecules in such composite films are demonstrated. The immobilization of DNA is accomplished by simple immersion of a thermally evaporated ODA film in the DNA solution at close to physiological pH. The entrapment of the DNA molecules in the cationic lipid film is dominated by attractive electrostatic interaction between the negatively charged phosphate backbone of the DNA molecules and the protonated amine molecules in the thermally evaporated film and has been quantified using quartz crystal microgravimetry (QCM). Fluorescence studies …

Cationic Surfactant Mediated Hybridization And Hydrophobization Of Dna Molecules At The Liquid/Liquid Interface And Their Phase Transfer, Murali Sastry, Ashavani Kumar, Mrunalini Pattarkine, Vidya Ramakrishnan, Krishna N. Ganesh

Faculty Works

Hybridization of complementary oligonucleotides mediated by a cationic surfactant at the water/hexane interface leads to hydrophobic, double-helical DNA which may be readily phase transferred to the organic phase and cast into thin films on solid substrates.

Aquaporin 5-Deficient Mouse Lungs Are Hyperresponsive To Cholinergic Stimulation, Carissa M. Krane, Christopher N. Fortner, Arthur R. Hand, Dennis W. Mcgraw, John N. Lorenz, Susan E. Wert, Jennifer E. Towne, Richard J. Paul, Jeffrey A. Whitsett, Anil G. Menon

Biology Faculty Publications

Although aquaporin 5 (AQP5) is the major water channel expressed in alveolar type I cells in the lung, its actual role in the lung is a matter of considerable speculation. By using immunohistochemical staining, we show that AQP5 expression in mouse lung is not restricted to type I cells, but is also detected in alveolar type II cells, and in tracheal and bronchial epithelium. Aqp5 knockout (Aqp5−/−) mice were used to analyze AQP5 function in pulmonary physiology. Compared with Aqp5+/+ mice, Aqp5−/− mice show a significantly increased concentration-dependent bronchoconstriction to intravenously administered Ach, as shown by …

Hawaiian Freshwater Polychaeta: A Potentially Substantial Trophic Component Of Stream Depositional Habitats, M. Eric Benbow, Albert J. Burky, Carl M. Way

Biology Faculty Publications

In this paper we report the widespread occurrence of large annelids (Polychaeta: Nereididae) in Hawaiian stream depositional habitats, drawing attention to the lack of knowledge of Hawaiian stream energetics. Specimens of Namalycastis sp. were collected from five Hawaiian Islands from May–July 1995 and from the island of Maui from July–December 1999 and April–July 2000. Most specimens collected were N. hawaiiensis, while two specimens of N. abiuma were collected from a single site on the island of Molokai. Specimens were collected from elevations ranging from 3–240 m; several collections were from sites above at least one major (> 15 m …

Hybridization Of Dna By Sequential Immobilization Of Oligonucleotides At The Air-Water Interface, Murali Sastry, Vidya Ramakrishnan, Mrunalini Pattarkine, Anand Gole, K. N. Ganesh

Faculty Works

The hybridization of DNA by sequential electrostatic and hydrogen-bonding immobilization of single-stranded complementary oligonucleotides at the air-water interface with cationic Langmuir monolayers is demonstrated. The complexation of the single-stranded DNA molecules with octadecylamine (ODA) Langmuir monolayers was followed in time by monitoring the pressure-area isotherms. A large (and slow) expansion of the ODA monolayer was observed during each stage of complexation in the following sequence: primary single-stranded DNA followed by complementary single-stranded DNA followed by the intercalator, ethidium bromide. Langmuir-Blodgett (LB) films of the ODA-DNA complex were formed on different substrates and characterized using quartz-crystal microgravimetry (QCM), Fourier transform infrared …

The Occurrence Of The Freshwater Clams, Musculium Partumeium (Say) And Pisidium Casertanum (Poli) (Bivalvia: Sphaeriidae), In The Hawaiian Islands, Albert J. Burky, Carl M. Way, Skippy Hau, M. Eric Benbow

Biology Faculty Publications

Sphaeriid clams were collected from an ancient, continuously cultivated taro pond complex at Ke‘anae Peninsula, Maui. Both species are known for adaptations for temporary pond habitats and could have been initially introduced by the earliest Hawaiian settlers transported with moist taro root stock from other areas of Oceania.

Electrically Mediated Plasmid Dna Delivery To Hepatocellular Carcinomas In Vivo, L. Heller, M. J. Jaroszeski, D. Coppola, C. Pottinger, R. Gilbert, Richard Heller

Bioelectrics Publications

Gene therapy by direct delivery of plasmid DNA has several advantages over viral gene transfer, but plasmid delivery is less efficient. In vivo electroporation has been used to enhance delivery of chemotherapeutic agents to tumors in both animal and human studies. Recently, this delivery technique has been extended to large molecules such as plasmid DNA. Here, the successful delivery of plasmids encoding reporter genes to rat hepatocellular carcinomas by in vivo electroporation is demonstrated.

Subcloning, Characterization And Sequencing Of The Streptomycin Resistance Gene Of A Multiple Antibiotic Resistance Transposon From Salmonella Typhi, Ramesvari Pararajasingam Pillai

Student Works (2000-2009)

Salmonella typhi S8 is resistant to ampicillin (Ac), chloramphenicol (Cm), cotrimoxazole (Ctm), streptomycin (Sm), and tetracycline (Tc). The Ac, Cm, Ctm. and Sm resistance traits have been shown to be mediated by a multiple antibiotic resistance transposon located on a large conjugative plasmid. Transposition of the transposon encoding Ac^R (ampicillin resistance), Cm^R Ctm^R and Sm^R designated TnX8, was done by Wong (1999) to a recipient replicon, plasmid pUB307. The Ac^R Cm^R, Ctm^R and Sm^R genes were subcloned into plasmid pKan by digesting the recombinant plasmid, des ignated pCL8 (pUB307::TnX8), with San. This subcloning strategy generated different recombinant pKans that harbour …

Genetic Diversity Of Salmonella Typhi, Satheesh Nair

Student Works (2000-2009)

Typhoid fever is a unique human septemic infection caused by Salmonella typhi. Typhoid fever is still an important public health problem in many developing countries. The continuing presence of the disease in endemic areas and emergence of multidrug- resistant strains in many developing countries as well as the increasing reports from developed countries has renewed the interest to better understand the epidemiology of typhoid fever and some aspects of its pathogenesis. Characterization of S. typhi strains in different epidemiological settings depends largely on the utility of highly precise molecular typing tools. A convenient, versatile and safe method for preparing bacterial …

Biotechnology In Western Australian Agriculture, Agriculture Western Australia

Grain and other field crops published reports

The Government of Western Australia believes that the strategic use of biotechnology and particularly gene technology has the potential to offer Western Australian agriculture significant economic benefits by improving the productivity and quality of products, as well as contributing to the development of sustainable farming systems.

Information contained in these binders is intended to give an overview of Biotechnology in relation to what is occurring in Australia, in Western Australia and specifically, at Agriculture Western Australia (AGWEST).

Biotechnology has been around for twenty-five years or more, but has only recently become a public issue in Australia - particularly in relation …

Genome Research And Traditional Intellectual Property Protection -- A Bad Fit?, Kate H. Murashige

RISK: Health, Safety & Environment (1990-2002)

Dr. Murashige addresses the need for a patent system more closely tailored to the needs of biotechnology. For example, the obviousness requirement may interfere with using patents to recoup high costs of work when it could arguably be done by researchers of ordinary skill.

Anion Induced Blue To Purple Transition In Bacteriorhodopsin, Mrunalini Pattarkine, Anil K. Singh

Faculty Works

Purple membrane (PM, λ" role="presentation">λmax" role="presentation">max 570 nm) of H. halobium on treatment with sulphuric acid changes its colour to blue (λ" role="presentation">λmax" role="presentation">max 608 nm). The purple chromophore can be regenerated from the blue chromophore by exogeneous addition of anions such as CI−" role="presentation">− and HPO42−" role="presentation">2−4. Chloride ion is found to be more effective than the dibasic phosphate ion in regenerating the purple chromophore. Nevertheless, one thing common to the anion regeneration is that both CI−" role="presentation">− and HPO42−" role="presentation">2−4 show marked pH effect. At pH 1.0 the efficiency of …

Metal-Resistance Genetically Engineered Bacteria, Sylvia Daunert, Donna Scott, Sridhar Ramanathan

KWRRI Research Reports

Bacterial-based electrochemical and optical sensing systems that respond in a highly selective and sensitive manner to antimonite and arsenite have been developed. This was accomplished by using genetically engineered bacteria bearing one of two plasmids constructed for our studies. The first plasmid, pBGD23, contains the operator/promoter region (O/P) and the gene of the ArsR protein from the ars operon upstream from the β-galactosidase gene. In the absence of antimonite/arsenite, ArsR binds to the 0/P site and prevents the transcription of the genes for ArsR and β-galactosidase, thus blocking expression of these proteins. When antimonite or arsenite is present in the …

Enhancer Trap Technique: A Novel Tool For Identification And Developmental Characterization Of Genes Of Drosophila, Amit Singh

Biology Faculty Publications

The classical technique of mutational screen for identification of genes controlling early development has now approached saturation. A new era in genetic identification and developmental characterization of genes in Drosophila has commenced with the advent of the enhancer trap technique. This technique involves mobilization of a P-lacZ vector to diverse chromosomal locations in the fruit fly genome to bring it under the regulation of developmentally expressed genes or their enhancer elements. The technique offers a strikingly elegant method of gaining entry into fruit fly genes.

An Improved Method For Chemical Devitellinization Of X-Gal Stained Drosophila Embryos, Amit Singh, Madhuri Kango-Singh, P. Sinha

Biology Faculty Publications

In Drosophila developmental biological studies, X-gal staining is commonly employed to study the spatio-temporal expression of the lacZ reporter gene in the transformed flies or their embryos. Study of the lacZ pattern in embryos often suffers from the lack of an efficient and high yieldirrg technique for devitellinization of X-gal stained embryos. Devitellinization techniques employed during antibody staining, in situ hybridization or embryonic cuticular preparations generally do not give satisfactory results when used for similar purpose in X-gal stained embryos. This results in the flaky appearance of the blue stain. We present here an improved chemical devitellinization technique which gives …