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In situ hybridization is an important technique for measuring the spatial expression patterns of mRNA in cells, tissues, and whole animals. However, mRNA levels cannot be compared across experiments using typical protocols. Here we present a semi-quantitative method to compare mRNA levels of a gene across multiple samples. This method yields an estimate of the error in the measurement to allow statistical...
The eye has been one of the most intensively studied organs in Drosophila. The wealth of knowledge about its development, as well as the reagents that have been developed, and the fact that the eye is dispensable for survival, also make the eye suitable for genetic interaction studies and genetic screens. This article provides a brief overview of the methods developed to image and probe eye development...
Drosophila oogenesis is an excellent system for the study of developmental cell biology. Active areas of research include stem cell maintenance, gamete development, pattern formation, cytoskeletal regulation, intercellular communication, intercellular transport, cell polarity, cell migration, cell death, morphogenesis, cell cycle control, and many more. The large size and relatively simple organization...
Here, I discuss how RNAi screening can be used effectively to uncover gene function. Specifically, I discuss the types of high-throughput assays that can be done in Drosophila cells and in vivo, RNAi reagent design and available reagent collections, automated screen pipelines, analysis of screen results, and approaches to RNAi results verification.
Circadian rhythms have a profound influence on most bodily functions: from metabolism to complex behaviors. They ensure that all these biological processes are optimized with the time-of-day. They are generated by endogenous molecular oscillators that have a period that closely, but not exactly, matches day length. These molecular clocks are synchronized by environmental cycles such as light intensity...
Modern biological research relies heavily on microscopic imaging. The advanced genetic toolkit of Drosophila makes it possible to label molecular and cellular components with unprecedented level of specificity necessitating the application of the most sophisticated imaging technologies. Imaging in Drosophila spans all scales from single molecules to the entire populations of adult organisms, from...
The TGF-β pathway is an evolutionarily conserved signal transduction module that mediates diverse biological processes in animals. In Drosophila, both the BMP and Activin branches are required for viability. Studies rooted in classical and molecular genetic approaches continue to uncover new developmental roles for TGF-β signaling. We present an overview of the secreted ligands, transmembrane receptors...
A vast diversity of biological systems, ranging from prokaryotes to multicellular organisms, show cell migration behavior. Many of the basic cellular and molecular concepts in cell migration apply to diverse model organisms. Drosophila, with its vast repertoire of tools for imaging and for manipulation, is one of the favorite organisms to study cell migration. Moreover, distinct Drosophila tissues...
In recent years the Drosophila heart has become an established model for many different aspects of human cardiac disease. This model has allowed identification of disease-causing mechanisms underlying congenital heart disease and cardiomyopathies and has permitted the study of underlying genetic, metabolic and age-related contributions to heart function. In this review we discuss methods currently...
The Mitogen-Activated Protein Kinase (MAPK) pathway represents one of the most conserved signaling cascades in multicellular organisms, since its cytoplasmic components can also be found in single-celled eukaryotic organisms such as yeast. With this broad view in mind, we can ask ourselves not only what are the seminal features and functions of the pathway in Drosophila, but also what have the studies...
The main Wnt ligand of Drosophila activates a conserved canonical signalling pathway to regulate a plethora of cellular activities during development, regeneration and nervous system function. Here I first describe experimental means of measuring and modulating Wingless signalling in Drosophila cell culture. Various reporters have been devised by placing TCF-binding sites or DNA fragments from known...
The Janus Kinase/Signal Transducers and Activators of Transcription (JAK/STAT) signaling pathway is one of a limited number of signaling cascades that is extensively utilized for many developmental and homeostatic functions. The JAK/STAT pathway is evolutionarily conserved from insects to mammals, with homologous transduction machinery in each. Yet the mammalian pathway is composed of multiple members...
•About 150 Drosophila cell lines from a variety of tissues sources exist in a community resource.•Extensive genomic and transcriptional data are available for many of these lines.•We review basic techniques for culture, transformation, and storage of cell lines.•We survey the wide range of applications for which Drosophila lines have been successfully used.
Planar cell polarity (PCP) is the polarity of epithelial cells in the plane orthogonal to the apical–basal axis, and is controlled by a partially defined signaling system [54,76]. PCP related signaling also plays roles in cell migration, tissue re-organization and stem cell differentiation during embryonic development, and later, in regeneration and repair [29,58]. Aberrant signaling has been linked...
The term autophagy refers to the engulfment and degradation of cytoplasmic components within the lysosome. This process can benefit cells and organisms by removing damaged, superfluous, or harmful cellular components, and by generating a supply of recycled macromolecules that can support biosynthesis or energy production. Recent interest in autophagy has been driven by its potential role in several...
Imaginal discs, especially the wing imaginal disc, are powerful model systems to study organ development. The traditional methods to analyze wing imaginal discs depend on the laborious and time-consuming dissection of larvae. “Omics”-based approaches, such as RNA-seq, ChIP-seq, proteomics and lipidomics, offer new opportunities for the systems-level investigation of organ development. However, it...
In this chapter we discuss methods that can be used to study apoptotic cell death in the Drosophila embryo, ovary, as well as in cultured cell lines. These methods assay various aspects of the cell death process, from mitochondrial changes to caspase activation and DNA cleavage. The assays are useful for examining apoptosis in normal development and in response to developmental perturbations and external...
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