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  JoVE Neuroscience

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  JoVE Immunology and Infection

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  JoVE Developmental Biology


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Pupa: An inactive stage between the larval and adult stages in the life cycle of insects.
 JoVE Neuroscience

Visualization of Proprioceptors in Drosophila Larvae and Pupae

1Department of Genetics and the Rappaport Institute for Research in the Medical Sciences, Rappaport Faculty of Medicine, Technion-Israel Institute of Technology

JoVE 3846

A method to immunostain and visualize chordotonal organs in larvae and pupae of Drosophila melanogaster is described.

 JoVE Biology

Primary Neuronal Cultures from the Brains of Late Stage Drosophila Pupae

1Department of Development and Cell Biology, Department of Anatomy and Neurobiology, University of California, Irvine (UCI)

JoVE 200

This video demonstrates the preparation of primary neuronal cultures from the brains of late stage Drosophila pupae. Views of live cultures show neurite outgrowth and imaging of calcium levels using Fura-2.

 JoVE Environment

Mass Production of Genetically Modified Aedes aegypti for Field Releases in Brazil

1Oxitec Ltd, 2Departamento de Parasitologia, Instituto de Ciências Biomédicas, Universidade de São Paulo, 3Departamento de Epidemiologia, Universidade de São Paulo, 4Moscamed Brasil, 5Deptartment of Zoology, University of Oxford, 6Instituto Nacional de Ciência e Tecnologia em Entomologia Molecular (INCT-EM)

JoVE 3579

To achieve population suppression of Aedes aegypti using the RIDL® (Release of Insects carrying a Dominant Lethal) system, large numbers of male mosquitoes need to be released. This requires the use of mass rearing techniques and technology to provide reliable systems to obtain the maximum number of high quality male mosquitoes.

 JoVE Biology

Ex vivo Culture of Drosophila Pupal Testis and Single Male Germ-line Cysts: Dissection, Imaging, and Pharmacological Treatment

1Fachbereich Biologie, Entwicklungsbiologie, Philipps-Universität Marburg, 2Institut für Molekularbiologie und Tumorforschung, Philipps-Universität Marburg

JoVE 51868

This protocol describes the dissection and cultivation of intact testes and germ-line cysts from Drosophila melanogaster pupae. This method allows microscopic observation of spermatogenesis ex vivo. Furthermore, we describe a pharmacological assay of the effect of inhibitors on specific stages of germ-cell development in pupal testes.

 JoVE Biology

Larval RNA Interference in the Red Flour Beetle, Tribolium castaneum

1Department of Biology, Miami University

JoVE 52059

RNA interference (RNAi)-based gene knockdown techniques are at the core of Tribolium research. Here, we provide an overview of our larval RNAi technique in Tribolium castaneum. Larval RNAi is a simple, but powerful technique that provides quick access to loss-of-function phenotypes, allowing researchers to study gene functions in diverse contexts.

 JoVE Biology

Chitosan/Interfering RNA Nanoparticle Mediated Gene Silencing in Disease Vector Mosquito Larvae

1Division of Biology, Kansas State University, 2Department of Medical and Molecular Genetics, Indiana University School of Medicine, 3Eck Institute for Global Health, University of Notre Dame, 4Department of Biological Sciences, University of Notre Dame, 5Department of Entomology, Kansas State University

JoVE 52523

Here we describe a procedure for inhibiting gene function in disease vector mosquitoes through the use of chitosan/interfering RNA nanoparticles that are ingested by larvae.

 JoVE Developmental Biology

Live-imaging of the Drosophila Pupal Eye

1Biology Department, Wesleyan University

JoVE 52120

This protocol presents an efficient method for imaging the live Drosophila pupal eye neuroepithelium. This method compensates for tissue movement and uneven topology, enhances visualization of cell boundaries through the use of multiple GFP-tagged junction proteins, and uses an easily-assembled imaging rig.

 JoVE Biology

Imaging Through the Pupal Case of Drosophila melanogaster

1Department of Biology, University of Miami

JoVE 51239

This paper demonstrates the use of a fast scanning confocal microscope to image cell behavior directly through the puparium. By leaving the pupal case intact, this method allows observation and measurement of dynamic cell processes at a stage of Drosophila development that is difficult to study directly.

 JoVE Neuroscience

Ex vivo Culturing of Whole, Developing Drosophila Brains

1National Institute of Neurological Disorders and Stroke, 2National Human Genome Research Institute, National Institutes of Health, Bethesda, MD

JoVE 4270

This article describes a method by which one can mimic in vivo development of the Drosophila mushroom body in an ex vivo culture system.

 JoVE Biology

An Experimental and Bioinformatics Protocol for RNA-seq Analyses of Photoperiodic Diapause in the Asian Tiger Mosquito, Aedes albopictus

1Department of Biology, Georgetown University, 2Insect Physiology Lab, EEOB, The Ohio State University

JoVE 51961

RNA-Seq analyses are becoming increasingly important for identifying the molecular underpinnings of adaptive traits in non-model organisms. Here, a protocol to identify differentially expressed genes between diapause and non-diapause Aedes albopictus mosquitoes is described, from mosquito rearing, to RNA sequencing and bioinformatics analyses of RNA-Seq data.

 JoVE Environment

Use of Chironomidae (Diptera) Surface-Floating Pupal Exuviae as a Rapid Bioassessment Protocol for Water Bodies

1Department of Entomology, University of Minnesota, 2Biology, Chemistry & Physics, and Mathematics Department, Northern State University, 3Environmental Analysis and Outcomes Division, Minnesota Pollution Control Agency, 4RMB Environmental Laboratories, Inc.

JoVE 52558

Rapid bioassessment protocols using benthic macroinvertebrates are often used to monitor and assess water quality. An efficient protocol involves collections of Chironomidae surface-floating pupal exuviae (SFPE). Here, techniques for field collection, laboratory processing, slide mounting, and identification of Chironomidae SFPE are described.

 JoVE Neuroscience

Dissection and Immunohistochemistry of Larval, Pupal and Adult Drosophila Retinas

1Department of Biology, New York University

JoVE 4347

The Drosophila retina is a crystal-like lattice composed of a small number of cell types that are generated in a stereotyped manner 1. Its amenability to sophisticated genetic analysis allows the study of complex developmental programs. This protocol describes dissections and immunohistochemistry of retinas at three discrete developmental stages, with a focus on photoreceptor differentiation.

 JoVE Behavior

Early Metamorphic Insertion Technology for Insect Flight Behavior Monitoring

1Department of Electrical and Computer Engineering, North Carolina State University

JoVE 50901

We present a novel surgical procedure to implant electrodes in Manduca sexta during its early metamorphic stages. This technique allows mechanically stable and electrically reliable coupling with the neuromuscular tissue to study flight neurophysiology dynamics. We also present a novel magnetic levitation platform for tethered studies of insect yaw.

 JoVE Biology

Live Cell Cycle Analysis of Drosophila Tissues using the Attune Acoustic Focusing Cytometer and Vybrant DyeCycle Violet DNA Stain

1Molecular, Cellular and Developmental Biology, University of Michigan

JoVE 50239

A protocol for cell cycle analysis of live Drosophila tissues using the Attune Acoustic Focusing Cytometer is described. This protocol simultaneously provides information about relative cell size, cell number, DNA content and cell type via lineage tracing or tissue specific expression of fluorescent proteins in vivo.

 JoVE Biology

Preparation of Developing and Adult Drosophila Brains and Retinae for Live Imaging

1Department of Physiology and Green Center for Systems Biology, University of Texas Southwestern Medical Center

JoVE 1936

This protocol describes three Drosophila preparations: 1) adult brain dissection, 2) adult retina dissection and 3) developing eye disc- brain complexes dissection. Emphasis is laid on special preparation techniques and conditions for live imaging, although all preparations can be used for fixed tissue immunohistochemistry.

 JoVE Biology

Simple Method for Fluorescence DNA In Situ Hybridization to Squashed Chromosomes

1Department of Biology, University of Rochester, 2W. M. Keck Science Department, Claremont McKenna, Pitzer, and Scripps Colleges

JoVE 52288

Here, we present a simple method for performing fluorescence DNA in situ hybridization (DNA ISH) to visualize repetitive heterochromatic sequences on slide-mounted chromosomes. The method requires minimal reagents and it is versatile for use with short or long probes, different tissues, and detection with fluorescence or non-fluorescence-based signals.

 JoVE Biology

Dissection and Mounting of Drosophila Pupal Eye Discs

1Department of Molecular, Cell and Developmental Biology (MCDB), University of California at Los Angeles (UCLA), 2Arizona College of Osteopathic Medicine (AZCOM), Midwestern University, 3MCDB, Broad Stem Cell Research Center, UCLA

JoVE 52315

The goal of this technique is to enable researchers to perform dissection, immunostaining and mounting of pupal eye discs from Drosophila melanogaster of any age.

 JoVE Immunology and Infection

Use of Galleria mellonella as a Model Organism to Study Legionella pneumophila Infection

1Center for Molecular Bacteriology and Infection, Imperial College London

JoVE 50964

The larva of the wax moth Galleria mellonella was recently established as an in vivo model to study Legionella pneumophila infection. Here, we demonstrate fundamental techniques to characterize the pathogenesis of Legionella in the larvae, including inoculation, measurement of bacterial virulence and replication as well as extraction and analysis of infected hemocytes.

 JoVE Biology

Imaging Centrosomes in Fly Testes

1Department of Biological Sciences, University of Toledo

JoVE 50938

Imaging of centrosomal proteins during Drosophila spermatogenesis is a powerful method to identify new proteins critical for centrosome biology as well as to elucidate the particular function of known players in this process.

 JoVE Immunology and Infection

An Introduction to Parasitic Wasps of Drosophila and the Antiparasite Immune Response

1Biology Department, The City College of New York, CUNY, 2The Graduate Center, The City University of New York

JoVE 3347

Parasitoid (parasitic) wasps constitute a major class of natural enemies of many insects including Drosophila melanogaster. We will introduce the techniques to propagate these parasites in Drosophila spp. and demonstrate how to analyze their effects on immune tissues of Drosophila larvae.

 JoVE Biology

Protocol for Mosquito Rearing (A. gambiae)

1Malaria Research Institute, Bloomberg School of Public Health, Johns Hopkins University

JoVE 221

This video illustrates the general techniques used to rear Anopheles gambiae in the laboratory. The methods for caring for laboratory mosquitoes are demonstrated through all stages of the organism's life cycle from larvae to pupae to blood-feeding adults.

 JoVE Immunology and Infection

The Insect Galleria mellonella as a Powerful Infection Model to Investigate Bacterial Pathogenesis

1INRA, Micalis UMR1319, France

JoVE 4392

Oral and intra haemocolic infection of larvae of the greater wax moth Galleria mellonella is described. This insect can be used to study virulence factors of entomopathogenic as well as mammalian opportunistic bacteria. Rearing of the insects, methods of infection and examples of in vivo analysis are described.

 JoVE Biology

Experimental Manipulation of Body Size to Estimate Morphological Scaling Relationships in Drosophila

1Department of Biology & Biochemistry, University of Houston, 2Department of Zoology, Michigan State University

JoVE 3162

Morphological scaling relationships capture and describe organismal shape. We present a method to measure morphological scaling relationships across the natural range of body sizes in fully metamorphic insects. Using a simple diet manipulation we increase the distribution of trait sizes, permitting the accurate description of how shape and size co-vary.

 JoVE Immunology and Infection

Systemic Bacterial Infection and Immune Defense Phenotypes in Drosophila Melanogaster

1Department of Entomology, Cornell University

JoVE 52613

Drosophila melanogaster is an outstanding model organism for studying innate immune systems and the physiological consequences of infection and disease. This protocol describes how to deliver robust and quantitatively repeatable bacterial infections to D. melanogaster, and how to subsequently measure infection severity and quantify the host immune response.

 JoVE Biology

The Tomato/GFP-FLP/FRT Method for Live Imaging of Mosaic Adult Drosophila Photoreceptor Cells

1Laboratory of Molecular Biology of the Cell, Ecole Normale Supérieure de Lyon, 2INSERM U744, Institut Pasteur de Lille, Université Lille-Nord de France, 3Howard Hughes Medical Institute, Laboratory of Apoptosis and Cancer, The Rockefeller University

JoVE 50610

The Tomato/GFP-FLP/FRT method involves visualizing mosaic photoreceptor cells in living Drosophila. It can be used to follow individual photoreceptor cell fates in the retina for days or weeks. This method is ideal for studies of retinal degeneration and neurodegenerative diseases or photoreceptor cell development.

 JoVE Bioengineering

Flying Insect Detection and Classification with Inexpensive Sensors

1Department of Computer Science and Engineering, University of California, Riverside, 2Department of Entomology, University of California, Riverside, 3Institute of Mathematics and Computer Sciences, University of São Paulo - USP, 4ISCA Technologies

JoVE 52111

We proposed a system that uses inexpensive, noninvasive pseudo-acoustic optical sensors to automatically and accurately detect, count, and classify flying insects based on their flying sound.

 JoVE Neuroscience

Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae

1Department of Biochemistry and Molecular Biology, The University of Texas MD Anderson Cancer Center, 2Scholars Academy/MARC Scholar, University of Houston-Downtown, 3Genes and Development Graduate Program, University of Texas Graduate School of Biomedical Sciences, 4Neuroscience Graduate Program, University of Texas Graduate School of Biomedical Sciences

JoVE 3837

In this article, we demonstrate assays to study thermal nociception in Drosophila larvae. One assay involves spatially-restricted (local) stimulation of thermal nociceptors1,2 while the second involves a wholesale (global) activation of most or all such neurons3. Together, these techniques allow visualization and quantification of the behavioral functions of Drosophila nociceptive sensory neurons.

 JoVE Biology

Electroantennographic Bioassay as a Screening Tool for Host Plant Volatiles

1Plant Mycotoxin Research, U.S. Department of Agriculture, Agricultural Research Service

JoVE 3931

A method to rapidly screen host plant volatiles by measurement of the electrophysiological response of adult navel orangeworm (Amyelois transitella) antennae to single components and blends via electroantennographic analysis is demonstrated.

 JoVE Biology

Toxicological Assays for Testing Effects of an Epigenetic Drug on Development, Fecundity and Survivorship of Malaria Mosquitoes

1Department of Entomology, Virginia Tech

JoVE 52041

A protocol is developed to examine the effects of an epigenetic drug DZNep on the development, fecundity and survivorship of mosquitoes. Here we describe procedures for the aqueous exposure of DZNep to immature mosquitoes and a blood-based exposure of DZNep to adult mosquitoes in addition to measuring SAH hydrolase inhibition.

 JoVE Neuroscience

Using Insect Electroantennogram Sensors on Autonomous Robots for Olfactory Searches

1UMR 7503, Laboratoire Lorrain de Recherche en Informatique et ses Applications (LORIA), Centre National de la Recherche Scientifique (CNRS), 2UMR 1392 iEES-Paris, Institut d'Ecologie et des Sciences de l'Environnement de Paris, 3Physics of Biological Systems, Institut Pasteur

JoVE 51704

We describe a protocol for using insect antennae in the form of electroantennograms (EAGs) on autonomous robots. Our experimental design allows stable recordings within a day and resolves individual odor patches up to 10 Hz. The efficiency of EAG sensors for olfactory searches is demonstrated in driving a robot toward an odor source.

 JoVE Immunology and Infection

Protocol for Production of a Genetic Cross of the Rodent Malaria Parasites

1National Institute of Allergy and Infectious Diseases, National Institutes of Health, 2School of Life Science, Xiamen University

JoVE 2365

Genetic crosses of rodent malaria parasites are performed by feeding two genetically distinct parasites to mosquitoes. Recombinant progeny are cloned from mouse blood after allowing mosquitoes to bite infected mice. This video shows how to produce genetic crosses of Plasmodium yoelii and is applicable to other rodent malaria parasites.

 JoVE Immunology and Infection

Procedures for Identifying Infectious Prions After Passage Through the Digestive System of an Avian Species

1Animal and Plant Health Inspection Service, Wildlife Services, National Wildlife Research Center, USDA

JoVE 50853

Scavengers have potential to translocate infectious transmissible spongiform encephalopathy prions in their feces to disease-free areas. We detail methods used to determine if mouse-adapted scrapie prions remain infectious after passage though the digestive tract of American crows (Corvus brachyrhynchos), a common consumer of dead animals.

 JoVE Biology

Physiological Recordings and RNA Sequencing of the Gustatory Appendages of the Yellow-fever Mosquito Aedes aegypti

1Agricultural Research Service, Henry A. Wallace Beltsville Agricultural Research Center, Plant Sciences Institute, Invasive Insect Biocontrol and Behavior Laboratory, United States Department of Agriculture

JoVE 52088

Using two methods to estimate gene expression in the major gustatory appendages of Aedes aegypti, we have identified the set of genes putatively underlying the neuronal responses to bitter and repulsive compounds, as determined by electrophysiological examination.

 JoVE Biology

In situ Protocol for Butterfly Pupal Wings Using Riboprobes

1Department of Biological Sciences, SUNY-University at Buffalo, 2Dept. Ecology and Evolutionary Biology, Yale University

JoVE 208

In order to examine gene expression in the pupal wing tissue of Bicyclus anynana, we present an optimized protocol for in situ hybridizations using riboprobes. We also provide guidelines for the further optimization of this protocol for use in pupal wings of other Lepidopteran species.

 Science Education: Essentials of Biology 1

Drosophila melanogaster Embryo and Larva Harvesting and Preparation

JoVE Science Education

Drosophila melanogaster embryos and larvae are easy to manipulate and develop rapidly by mechanisms that are analogous to other organisms, including mammals. For these reasons, many researchers utilize fly embryos and larvae to answer questions in diverse fields ranging from behavioral to developmental biology. Prior to experimentation, however, the embryos and larvae must first be collected. This video will first demonstrate how "egg-laying cups" are used to collect Drosophila embryos on agar plates. The harvest and dechorionation of embryos will then be described. Next, the video will demonstrate how to identify and manipulate Drosophila in one of the three larval stages that follow the embryo stage. Finally, examples of some of the ways in which fly embryos and larvae are used in biological research are provided.

 Science Education: Essentials of Biology 1

Drosophila Maintenance

JoVE Science Education

Drosophila melanogaster, commonly known as fruit flies, are a frequently used model organism for life science research. Although starting a collection of these critters may seem as easy as leaving a banana on your kitchen counter for too long, a productive fly colony in the lab requires careful husbandry and maintenance.

This video demonstrates the necessary steps for maintaining a healthy fly stock. The overview begins with the preparation and storage of the yeast and sugar-containing media on which flies feed. Next, the vessels most commonly used for housing Drosophila are shown, as well as how and when to move flies between these containers. Finally, the presentation also includes examples of the ways in which housing and feeding conditions are manipulated for biological experiments.

 JoVE Biology

Operant Learning of Drosophila at the Torque Meter

1Department of Neurobiology, Free University of Berlin

JoVE 731

Measuring the yaw torque of tethered Drosophila with the torque meter allows the neuroscientist exquisite control of the stimulus situation of the experimental animal. Together with the unique genetic tools available in the fruit fly, this paradigm is used for a wide variety of neurobiological research.

 JoVE Biology

Mesoscopic Fluorescence Tomography for In-vivo Imaging of Developing Drosophila

1Center for Systems Biology, Massachusetts General Hospital, 2Institute for Biological and Medical Imaging (IBMI), Technical University of Munich and Helmholtz Center Munich, 3Department of Genetics, Harvard Medical School and Howard Hughes Medical Institute

JoVE 1510

Mesoscopic fluorescence tomography operates beyond the penetration limits of tissue-sectioning fluorescence microscopy. The technique is based on multi-projection illumination and a photon transport description. We demonstrate in-vivo whole-body 3D visualization of the morphogenesis of GFP-expressing wing imaginal discs in Drosophila melanogaster.

 JoVE Biology

An Optimized Protocol for Rearing Fopius arisanus, a Parasitoid of Tephritid Fruit Flies

1Agricultural Research Service, US Dept. of Agriculture, US Pacific Basin Agricultural Research Center

JoVE 2901

Fopius arisanus is an egg-larval parasitoid of Tephritid fruit flies that is successfully used in biological control of these important tropical pests. We describe here an optimized protocol for rearing F. arisanus on a small scale using readily available materials.

 JoVE Biology

Quantitative Comparison of cis-Regulatory Element (CRE) Activities in Transgenic Drosophila melanogaster

1Department of Biology, University of Dayton, 2Department of Biology, Center for Tissue Regeneration and Engineering at Dayton, University of Dayton

JoVE 3395

Phenotypic variation for traits can result from mutations in cis-regulatory element (CRE) sequences that control gene expression patterns. Methods derived for use in Drosophila melanogaster can quantitatively compare the levels of spatial and temporal patterns of gene expression mediated by modified or naturally occurring CRE variants.

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