共查询到20条相似文献,搜索用时 31 毫秒
1.
Karen G. Hales Christopher A. Korey Amanda M. Larracuente David M. Roberts 《Genetics》2015,201(3):815-842
Fruit flies of the genus Drosophila have been an attractive and effective genetic model organism since Thomas Hunt Morgan and colleagues made seminal discoveries with them a century ago. Work with Drosophila has enabled dramatic advances in cell and developmental biology, neurobiology and behavior, molecular biology, evolutionary and population genetics, and other fields. With more tissue types and observable behaviors than in other short-generation model organisms, and with vast genome data available for many species within the genus, the fly’s tractable complexity will continue to enable exciting opportunities to explore mechanisms of complex developmental programs, behaviors, and broader evolutionary questions. This primer describes the organism’s natural history, the features of sequenced genomes within the genus, the wide range of available genetic tools and online resources, the types of biological questions Drosophila can help address, and historical milestones. 相似文献
2.
Mark B Keroles Sonya K Dusseault Chuchu Liu Masood R Mohammed Christy M Vadakkan Jessica H Amiel Samantha N Abel Elena R Bensoussan Benjamin L Russell James Baker 《Journal of visualized experiments : JoVE》2014,(83)
The longstanding use of Drosophila as a model for cell and developmental biology has yielded an array of tools. Together, these techniques have enabled analysis of cell and developmental biology from a variety of methodological angles. Live imaging is an emerging method for observing dynamic cell processes, such as cell division or cell motility. Having isolated mutations in uncharacterized putative cell cycle proteins it became essential to observe mitosis in situ using live imaging. Most live imaging studies in Drosophila have focused on the embryonic stages that are accessible to manipulation and observation because of their small size and optical clarity. However, in these stages the cell cycle is unusual in that it lacks one or both of the gap phases. By contrast, cells of the pupal wing of Drosophila have a typical cell cycle and undergo a period of rapid mitosis spanning about 20 hr of pupal development. It is easy to identify and isolate pupae of the appropriate stage to catch mitosis in situ. Mounting intact pupae provided the best combination of tractability and durability during imaging, allowing experiments to run for several hours with minimal impact on cell and animal viability. The method allows observation of features as small as, or smaller than, fly chromosomes. Adjustment of microscope settings and the details of mounting, allowed extension of the preparation to visualize membrane dynamics of adjacent cells and fluorescently labeled proteins such as tubulin. This method works for all tested fluorescent proteins and can capture submicron scale features over a variety of time scales. While limited to the outer 20 µm of the pupa with a conventional confocal microscope, this approach to observing protein and cellular dynamics in pupal tissues in vivo may be generally useful in the study of cell and developmental biology in these tissues. 相似文献
3.
Mazen S. Habayeb Gabriela Casanova Shannon Albright 《Journal of invertebrate pathology》2009,101(1):29-33
The biology of the Drosophila viruses has not been intensely investigated. Here we have investigated the biology of the Nora virus, a persistent Drosophila virus. We find that injected Nora virus is able to replicate in the files, reaching a high titer that is maintained in the next generation. There is a remarkable variation in the viral loads of individual flies in persistently infected stocks; the titers can differ by three orders of magnitude. The Nora virus is mainly found in the intestine of infected flies, and the histology of these infected intestines show increased vacuolization. The virus is excreted in the feces and is horizontally transmitted. The Nora virus infection has a very mild effect on the longevity of the flies, and no significant effect on the number of eggs laid and the percent of eggs that develop to adults. 相似文献
4.
Changes in dopamine (DA) signaling have been implicated in a number of human neurologic
and psychiatric disorders. Similarly, defects in DA signaling in the fruit fly,
Drosophila melanogaster, have also been associated with several
behavioral defects. As most genes involved in DA synthesis, transport, secretion, and
signaling are conserved between species, Drosophila is a powerful genetic
model organism to study the regulation of DA signaling in vivo. In this
review, we will provide an overview of the genes and drugs that regulate DA biology in
Drosophila. Furthermore, we will discuss the behavioral paradigms that
are regulated by DA signaling in flies. By analyzing the genes and neuronal circuits that
govern such behaviors using sophisticated genetic, pharmacologic, electrophysiologic, and
imaging approaches in Drosophila, we will likely gain a better
understanding about how this neuromodulator regulates motor tasks and cognition in
humans. 相似文献
5.
Poojitha Sitaram Sarah Grace Hainline Laura Anne Lee 《Journal of visualized experiments : JoVE》2014,(83)
Drosophila melanogaster is a powerful model system that has been widely used to elucidate a variety of biological processes. For example, studies of both the female and male germ lines of Drosophila have contributed greatly to the current understanding of meiosis as well as stem cell biology. Excellent protocols are available in the literature for the isolation and imaging of Drosophila ovaries and testes3-12. Herein, methods for the dissection and preparation of Drosophila testes for microscopic analysis are described with an accompanying video demonstration. A protocol for isolating testes from the abdomen of adult males and preparing slides of live tissue for analysis by phase-contrast microscopy as well as a protocol for fixing and immunostaining testes for analysis by fluorescence microscopy are presented. These techniques can be applied in the characterization of Drosophila mutants that exhibit defects in spermatogenesis as well as in the visualization of subcellular localizations of proteins. 相似文献
6.
Ranjini Prithviraj Svetlana Trunova Edward Giniger 《Journal of visualized experiments : JoVE》2012,(65)
We describe a method for ex vivo culturing of whole Drosophila brains. This can be used as a counterpoint to chronic genetic manipulations for investigating the cell biology and development of central brain structures by allowing acute pharmacological interventions and live imaging of cellular processes. As an example of the technique, prior work from our lab1 has shown that a previously unrecognized subcellular compartment lies between the axonal and somatodendritic compartments of axons of the Drosophila central brain. The development of this compartment, referred to as the axon initial segment (AIS)2, was shown genetically to depend on the neuron-specific cyclin-dependent kinase, Cdk5. We show here that ex vivo treatment of wild-type Drosophila larval brains with the Cdk5-specific pharmacological inhibitors roscovitine and olomoucine3 causes acute changes in actin organization, and in localization of the cell-surface protein Fasciclin 2, that mimic the changes seen in mutants that lack Cdk5 activity genetically.A second example of the ex vivo culture technique is provided for remodeling of the connections of embryonic mushroom body (MB) gamma neurons during metamorphosis from larva to adult. The mushroom body is the center of olfactory learning and memory in the fly4, and these gamma neurons prune their axonal and dendritic branches during pupal development and then re-extend branches at a later timepoint to establish the adult innervation pattern5. Pruning of these neurons of the MB has been shown to occur via local degeneration of neurite branches6, by a mechanism that is triggered by ecdysone, a steroid hormone, acting at the ecdysone receptor B17, and that is dependent on the activity of the ubiquitin-proteasome system6. Our method of ex vivo culturing can be used to interrogate further the mechanism of developmental remodeling. We found that in the ex vivo culture setting, gamma neurons of the MB recapitulated the process of developmental pruning with a time course similar to that in vivo. It was essential, however, to wait until 1.5 hours after puparium formation before explanting the tissue in order for the cells to commit irreversibly to metamorphosis; dissection of animals at the onset of pupariation led to little or no metamorphosis in culture. Thus, with appropriate modification, the ex vivo culture approach can be applied to study dynamic as well as steady state aspects of central brain biology. 相似文献
7.
Sarah E. Allen Gabriel T. Koreman Ankita Sarkar Bei Wang Mariana F. Wolfner Chun Han 《PLoS biology》2021,19(1)
Mosaic animals have provided the platform for many fundamental discoveries in developmental biology, cell biology, and other fields. Techniques to produce mosaic animals by mitotic recombination have been extensively developed in Drosophila melanogaster but are less common for other laboratory organisms. Here, we report mosaic analysis by gRNA-induced crossing-over (MAGIC), a new technique for generating mosaic animals based on DNA double-strand breaks produced by CRISPR/Cas9. MAGIC efficiently produces mosaic clones in both somatic tissues and the germline of Drosophila. Further, by developing a MAGIC toolkit for 1 chromosome arm, we demonstrate the method’s application in characterizing gene function in neural development and in generating fluorescently marked clones in wild-derived Drosophila strains. Eliminating the need to introduce recombinase-recognition sites into the genome, this simple and versatile system simplifies mosaic analysis in Drosophila and can in principle be applied in any organism that is compatible with CRISPR/Cas9.Analysis of mosaic animals has been crucial in developmental and cell biology; this study describes a versatile, simple, and likely widely-applicable technique, MAGIC (mosaic analysis by gRNA-induced crossing-over), for generating mosaic animals based on DNA double-strand breaks produced by CRISPR/Cas9. 相似文献
8.
Jacob M. Sawyer 《Developmental biology》2010,341(1):5-5188
Biologists have long recognized that dramatic bending of a cell sheet may be driven by even modest shrinking of the apical sides of cells. Cell shape changes and tissue movements like these are at the core of many of the morphogenetic movements that shape animal form during development, driving processes such as gastrulation, tube formation, and neurulation. The mechanisms of such cell shape changes must integrate developmental patterning information in order to spatially and temporally control force production—issues that touch on fundamental aspects of both cell and developmental biology and on birth defects research. How does developmental patterning regulate force-producing mechanisms, and what roles do such mechanisms play in development? Work on apical constriction from multiple systems including Drosophila, Caenorhabditis elegans, sea urchin, Xenopus, chick, and mouse has begun to illuminate these issues. Here, we review this effort to explore the diversity of mechanisms of apical constriction, the diversity of roles that apical constriction plays in development, and the common themes that emerge from comparing systems. 相似文献
9.
The Drosophila melanogaster embryo has been widely utilized as a model for genetics and developmental biology due to its small size, short generation time, and large brood size. Information on embryonic metabolism during developmental progression is important for further understanding the mechanisms of Drosophila embryogenesis. Therefore, the aim of this study is to assess the changes in embryos’ metabolome that occur at different stages of the Drosophila embryonic development. Time course samples of Drosophila embryos were subjected to GC/MS-based metabolome analysis for profiling of low molecular weight hydrophilic metabolites, including sugars, amino acids, and organic acids. The results showed that the metabolic profiles of Drosophila embryo varied during the course of development and there was a strong correlation between the metabolome and different embryonic stages. Using the metabolome information, we were able to establish a prediction model for developmental stages of embryos starting from their high-resolution quantitative metabolite composition. Among the important metabolites revealed from our model, we suggest that different amino acids appear to play distinct roles in different developmental stages and an appropriate balance in trehalose-glucose ratio is crucial to supply the carbohydrate source for the development of Drosophila embryo. 相似文献
10.
11.
Pierre Dourlen Clemence Levet Alexandre Mejat Alexis Gambis Bertrand Mollereau 《Journal of visualized experiments : JoVE》2013,(79)
The Drosophila eye is widely used as a model for studies of development and neuronal degeneration. With the powerful mitotic recombination technique, elegant genetic screens based on clonal analysis have led to the identification of signaling pathways involved in eye development and photoreceptor (PR) differentiation at larval stages. We describe here the Tomato/GFP-FLP/FRT method, which can be used for rapid clonal analysis in the eye of living adult Drosophila. Fluorescent photoreceptor cells are imaged with the cornea neutralization technique, on retinas with mosaic clones generated by flipase-mediated recombination. This method has several major advantages over classical histological sectioning of the retina: it can be used for high-throughput screening and has proved an effective method for identifying the factors regulating PR survival and function. It can be used for kinetic analyses of PR degeneration in the same living animal over several weeks, to demonstrate the requirement for specific genes for PR survival or function in the adult fly. This method is also useful for addressing cell autonomy issues in developmental mutants, such as those in which the establishment of planar cell polarity is affected. 相似文献
12.
The Drosophila neuromuscular junction (NMJ) is an established model system used for the study of synaptic development and plasticity. The widespread use of the Drosophila motor system is due to its high accessibility. It can be analyzed with single-cell resolution. There are 30 muscles per hemisegment whose arrangement within the peripheral body wall are known. A total of 31 motor neurons attach to these muscles in a pattern that has high fidelity. Using molecular biology and genetics, one can create transgenic animals or mutants. Then, one can study the developmental consequences on the morphology and function of the NMJ. Immunohistochemistry can be used to clearly image the components of the NMJ. In this article, we demonstrate how to use antibody staining to visualize the Drosophila larval NMJ. 相似文献
13.
Drosophila, a dipteran insect, has been found to be the best biological model for different kinds of studies. D melanogaster was first described by Meigen in 1830, is most extensively studied species of the genus Drosophila and a number of investigations employing this species have been documented in areas such as genetics, behaviour, evolution, development, molecular biology, ecology, population biology, etc. Besides D. melanogaster, a number of other species of the genus Drosophila have also been used for different kinds of investigations. Among these, D. ananassae, a cosmopolitan and domestic species endowed with several unusual genetic features, is noteworthy. Described for the first time from Indonesia (Doleschall 1858), this species is commonly distributed in India. Extensive research work on D. ananassae has been done by numerous researchers pertaining to cytology, genetics, mutagenesis, gene mapping, crossing-over in both sexes, population and evolutionary genetics, behaviour genetics, ecological genetics, sexual isolation, fluctuating asymmetry, trade-offs etc. Genome of D. ananassae has also been sequenced. The status of research on D. ananassae at global level is briefly described in this review. Bibliography on this species from different countries worldwide reveals that maximum contribution is from India. 相似文献
14.
Daniel J. Coleman Meenakshi Venkatesan Staci P. Williamson John J. Naleway 《Analytical biochemistry》2010,399(1):7-12
A simple and reliable continuous assay for measurement of α-mannosidase activity is described and demonstrated for analysis with two recombinant human enzymes using the new substrate resorufin α-d-mannopyranoside (Res-Man). The product of enzyme reaction, resorufin, exhibits fluorescence emission at 585 nm with excitation at 571 nm and has a pKa of 5.8, allowing continuous measurement of fluorescence turnover at or near physiological pH values for human lysosomal and Drosophila Golgi α-mannosidases. The assay performed using recombinant Drosophila Golgi α-mannosidase (dGMII) has been shown to give the kinetic parameters Km of 200 μM and Vmax of 11 nmol/min per nmol dGMII. Methods for performing the assay using several concentrations of the known α-mannosidase inhibitor swainsonine are also presented, demonstrating a potential for use of the assay as a simple method for high-throughput screening of inhibitors potentially useful in cancer treatment. 相似文献
15.
Alexey Veraksa 《Journal of Proteomics》2010,73(11):2158-2170
In the past decade, improvements in genome annotation, protein fractionation methods and mass spectrometry instrumentation resulted in rapid growth of Drosophila proteomics. This review presents the current status of proteomics research in the fly. Areas that have seen major advances in recent years include efforts to map and catalog the Drosophila proteome and high-throughput as well as targeted studies to analyze protein–protein interactions and post-translational modifications. Stable isotope labeling of flies and other applications of quantitative proteomics have opened up new possibilities for functional analyses. It is clear that proteomics is becoming an indispensable tool in Drosophila systems biology research that adds a unique dimension to studying gene function. 相似文献
16.
Paul Lasko 《Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms》2013,1829(8):810-816
DEAD-box helicases related to the Drosophila protein Vasa (also known as Ddx4) are found throughout the animal kingdom. They have been linked to numerous processes in gametogenesis, germ cell specification, and stem cell biology, and alterations in Vasa expression are associated with malignancy of tumor cells and with some human male infertility syndromes. Experimental results indicating how Vasa contributes to all these different cellular and developmental processes are discussed, using examples from planarians, Caenorhabditis elegans, Drosophila, sea urchin, zebrafish, Xenopus, mouse, and human. Molecular, cellular, and developmental functions of Vasa and its orthologs are reviewed in this article. Evidence linking Vasa to translational regulation, to biogenesis of small RNAs, and to chromosome condensation is examined. Finally, potential overlapping functions between Vasa and related DEAD-box helicases (Belle, or Ddx3, and DEADSouth, or Ddx25) are explored. This article is part of a Special Issue entitled: The biology of RNA helicases — Modulation for life. 相似文献
17.
18.
Adam Kleinschmit Masahiko Takemura Katsufumi Dejima Pui Yee Choi Hiroshi Nakato 《The Journal of biological chemistry》2013,288(7):5081-5089
Heparan sulfate proteoglycans regulate various physiological and developmental processes through interactions with a number of protein ligands. Heparan sulfate (HS)-ligand binding depends on the amount and patterns of sulfate groups on HS, which are controlled by various HS sulfotransferases in the Golgi apparatus as well as extracellular 6-O-endosulfatases called “Sulfs.” Sulfs are a family of secreted molecules that specifically remove 6-O-sulfate groups within the highly sulfated regions on HS. Vertebrate Sulfs promote Wnt signaling, whereas the only Drosophila homologue of Sulfs, Sulf1, negatively regulates Wingless (Wg) signaling. To understand the molecular mechanism for the negative regulation of Wg signaling by Sulf1, we studied the effects of Sulf1 on HS-Wg interaction and Wg stability. Sulf1 overexpression strongly inhibited the binding of Wg to Dally, a potential target heparan sulfate proteoglycan of Sulf1. This effect of Drosophila Sulf1 on the HS-Wg interaction is similar to that of vertebrate Sulfs. Using in vitro, in vivo, and ex vivo systems, we show that Sulf1 reduces extracellular Wg protein levels, at least partly by facilitating Wg degradation. In addition, expression of human Sulf1 in the Drosophila wing disc lowers the levels of extracellular Wg protein, as observed for Drosophila Sulf1. Our study demonstrates that vertebrate and Drosophila Sulfs have an intrinsically similar activity and that the function of Sulfs in the fate of Wnt/Wg ligands is context-dependent. 相似文献
19.
Recording and describing animal ‘monsters’ collected in the field can still contribute to progress in developmental biology despite the uncontrolled conditions the specimen experienced throughout development. Comparison with model organisms and a sound phylogenetic analysis may offer a tentative explanation for the underlying developmental mechanism and suggest new targets for experimental studies. We describe a female specimen of the anthomyiid fly Hydrophoria sp. with an ectopic macrochaeta in the left eye and suggest tentative interpretations, including one in terms of a local expression, or derepression, of a proneural gene. The anthomyiid lineage has been estimated to have split ca. 65 million years ago from the dipteran clade containing Drosophila and ca. 140 million years ago from the clade containing Megaselia. 相似文献
20.
The Drosophila lymph gland (LG) is a model system for studying hematopoiesis and blood cell homeostasis. Here, we investigated the patterns of division and differentiation of pro-hemocytes in normal developmental conditions and response to wasp parasitism, by combining lineage analyses and molecular markers for each of the three hemocyte types. Our results show that the embryonic LG contains primordial hematopoietic cells which actively divide to give rise to a pool of pro-hemocytes. We found no evidence for the existence of bona fide stem cells and rather suggest that Drosophila pro-hemocytes are regulated as a group of cells, rather than individual stem cells. The fate-restriction of plasmatocyte and crystal cell progenitors occurs between the end of embryogenesis and the end of the first larval instar, while Notch activity is required for the differentiation of crystal cells in third instar larvae only. Upon parasitism, lamellocyte differentiation prevents crystal cell differentiation and lowers plasmatocyte production. We also found that a new population of intermediate progenitors appears at the onset of hemocyte differentiation and accounts for the increasing number of differentiated hemocytes in the third larval instar. These findings provide a new framework to identify parameters of developmental plasticity of the Drosophila lymph gland and hemocyte homeostasis in physiological conditions and in response to immunological cues. 相似文献