Methods for Live Microscopy of Drosophila Spermatocytes

Matthew S Savoian, University of Cambridge, Cambridge, UK
David M Glover, University of Cambridge, Cambridge, UK

Published online: September 2008

DOI: 10.1002/9780470015902.a0020869

Drosophila is a genetically tractable system well suited for cell cycle investigations. The combination of live cell imaging and the variety of different cell types available, in particular, the easily cultured, exceptionally large and flat, primary spermatocyte, offer powerful tools for studying dynamic cell division events.

Keywords: live cell imaging; mitosis; meiosis; light microscopy; GFP

Figure 1. Primary spermatocyte culture system. (a) The aluminium slide and attached coverslip used as a culture chamber. (b) Testes (arrowheads) isolated from a pharate adult. (c) The primary culture of individual spermatocytes ready for live cell observation. Two cells entering division as revealed by their nuclear morphology (asterisks) are visible in the field. Bar is 20 m.
Figure 2. An example of a multidimensional data set for a spermatocyte with GFP-tagged microtubules. (a) The single, centre-most optical section as recorded by DIC imaging. Note the well-resolved and equatorially positioned metaphase chromosomes (arrowheads). (b) Six consecutive GFP fluorescence optical sections have been acquired through the entire volume of the cell and superimposed into a single image revealing all of the spindle's microtubules. (c) Images from (a) and (b) have been combined to reveal the spatial relationship between the spindle, chromosomes (arrowheads) and the cell's outer cortical boundaries. Bar is 10 m.
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 References
    Church K and Lin HP (1985) Kinetochore microtubules and chromosome movement during prometaphase in Drosophila melanogaster spermatocytes studied in life and with the electron microscope. Chromosoma 92: 273–282.
    Endow SA and Komma DJ (1996) Centrosome and spindle function of the Drosophila Ncd microtubule motor visualized in live embryos using Ncd-GFP fusion proteins. Journal of Cell Science 109: 2429–2442.
    Endow SA and Komma DJ (1997) Spindle dynamics during meiosis in Drosophila oocytes. Journal of Cell Biology 137: 1321–1336.
    Fleming SL and Rieder CL (2003) Flattening Drosophila cells for high-resolution light microscopic studies of mitosis in vitro. Cell Motility and the Cytoskeleton 56: 141–146.
    Goshima G and Vale RD (2003) The roles of microtubule-based motor proteins in mitosis: comprehensive RNAi analysis in the Drosophila S2 cell line. Journal of Cell Biology 162: 1003–1016.
    Goshima G, Wollman R, Goodwin SS et al. (2007) Genes required for mitotic spindle assembly in Drosophila S2 cells. Science 316: 417–421.
    Inoue YH, Savoian MS, Suzuki T et al. (2004) Mutations in orbit/mast reveal that the central spindle is comprised of two microtubule populations, those that initiate cleavage and those that propagate furrow ingression. Journal of Cell Biology 166: 49–60.
    Rebollo E and Gonzalez C (2000) Visualizing the spindle checkpoint in Drosophila spermatocytes. EMBO Reports 1: 65–70.
    Rebollo E, Llamazares S, Reina J and Gonzalez C (2004) Contribution of noncentrosomal microtubules to spindle assembly in Drosophila spermatocytes. PLoS Biology 2: 54–64.
    Savoian MS, Goldberg ML and Rieder CL (2000) The rate of poleward chromosome motion is attenuated in Drosophila zw10 and rod mutants. Nature Cell Biology 2: 948–952.
    Savoian MS and Rieder CL (2002) Mitosis in primary cultures of Drosophila melanogaster larval neuroblasts. Journal of Cell Science 115: 3061–3072.
    Siller KH, Serr M, Steward R, Hays TS and Doe CQ (2005) Live imaging of Drosophila brain neuroblasts reveals a role for lis1/dynactin in spindle assembly and mitotic checkpoint control. Molecular Biology of the Cell 16: 5127–5140.
    Theurkauf WE (1994) Premature microtubule-dependent cytoplasmic streaming in cappuccino and spire mutant oocytes. Science 265: 2093–2096.
 Further Reading
    book Hazelrigg T (2000) "GFP and other reporters". In: Sullivan W, Ashburner M and Hawley RS (eds) Drosophila Protocols, pp. 313–343. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press.
    Khodjakov A and Rieder CL (2006) Imaging the division process in living tissue culture cells. Methods 38: 2–16.
    book Matthies HJG, Clarkson M, Saint RB, Namba R and Hawley RS (2000) "Analysis of meiosis in fixed and live oocytes by light microscopy". In: Sullivan W, Ashburner M and Hawley RS (eds) Drosophila Protocols, pp. 67–85. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press.
    Rebollo E and Gonzalez C (2004) Time-lapse imaging of male meiosis by phase-contrast and fluorescence microscopy. Methods in Molecular Biology 247: 77–87.

Related Sub-Topics:

Cell Cycle | Techniques in Cell Biology | Optical Microscopy
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Article Title: Methods for Live Microscopy of Drosophila Spermatocytes
 
How to Cite close
Savoian, Matthew S, and Glover, David M(Sep 2008) Methods for Live Microscopy of Drosophila Spermatocytes. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0020869]