Abstract
Drosophila melanogaster has been studied for a century as a genetic model to understand recombination, chromosome segregation, and the basic rules of inheritance. However, it has only been about 25 years since the events that occur during nuclear envelope breakdown, spindle assembly, and chromosome orientation during D. melanogaster female meiosis I were first visualized by fixed cytological methods (Theurkauf and Hawley, J Cell Biol 116:1167–1180, 1992). Although these fixed cytological studies revealed many important details about the events that occur during meiosis I, they failed to elucidate the timing or order of these events. The development of protocols for live imaging of meiotic events within the oocyte has enabled collection of real-time information on the kinetics and dynamics of spindle assembly, as well as the behavior of chromosomes during prometaphase I. Here, we describe a method to visualize spindle assembly and chromosome movement during meiosis I by injecting fluorescent dyes to label microtubules and DNA into stage 12–14 oocytes. This method enables the events during Drosophila female meiosis I, such as spindle assembly and chromosome movement, to be observed in vivo, regardless of genetic background, with exceptional spatial and temporal resolution.
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References
King RC (1970) Ovarian development in Drosophila melanogaster. Academic, New York, NY
Theurkauf WE, Hawley RS (1992) Meiotic spindle assembly in Drosophila females: behavior of nonexchange chromosomes and the effects of mutations in the nod kinesin-like protein. J Cell Biol 116:1167–1180
Gilliland WD, Hughes SE, Vietti DR, Hawley RS (2009) Congression of achiasmate chromosomes to the metaphase plate in Drosophila melanogaster oocytes. Dev Biol 325:122–128
Hughes SE, Gilliland WD, Cotitta JL, Takeo S, Collins KA et al (2009) Heterochromatic threads connect oscillating chromosomes during prometaphase I in Drosophila oocytes. PLoS Genet 5:e1000348
Matthies HJ, McDonald HB, Goldstein LS, Theurkauf WE (1996) Anastral meiotic spindle morphogenesis: role of the non-claret disjunctional kinesin-like protein. J Cell Biol 134:455–464
Colombie N, Cullen CF, Brittle AL, Jang JK, Earnshaw WC et al (2008) Dual roles of Incenp crucial to the assembly of the acentrosomal metaphase spindle in female meiosis. Development 135:3239–3246
Endow SA, Komma DJ (1997) Spindle dynamics during meiosis in Drosophila oocytes. J Cell Biol 137:1321–1336
Colombie N, Gluszek AA, Meireles AM, Ohkura H (2013) Meiosis-specific stable binding of augmin to acentrosomal spindle poles promotes biased microtubule assembly in oocytes. PLoS Genet 9:e1003562
Hughes SE, Beeler JS, Seat A, Slaughter BD, Unruh JR et al (2011) Gamma-tubulin is required for bipolar spindle assembly and for proper kinetochore microtubule attachments during prometaphase I in Drosophila oocytes. PLoS Genet 7:e1002209
Gilliland WD, Hughes SE, Cotitta JL, Takeo S, **ang Y et al (2007) The multiple roles of Mps1 in Drosophila female meiosis. PLoS Genet 3:e113
**ang Y, Takeo S, Florens L, Hughes SE, Huo LJ et al (2007) The inhibition of polo kinase by matrimony maintains G2 arrest in the meiotic cell cycle. PLoS Biol 5:e323
Endow SA, Hallen MA (2011) Anastral spindle assembly and gamma-tubulin in Drosophila oocytes. BMC Cell Biol 12:1
Liang ZY, Hallen MA, Endow SA (2009) Mature Drosophila meiosis I spindles comprise microtubules of mixed polarity. Curr Biol 19:163–168
Davis I (2000) Visualizing fluorescence in Drosophila – optimal detection in thick specimens. In: Allan VJ (ed) Protein localization by fluorescence microscopy: a practical approach. Oxford University Press, New York, NY
Miller DF, Holtzman SL, Kaufman TC (2002) Customized microinjection glass capillary needles for P-element transformations in Drosophila melanogaster. Biotechniques 33:366–367, 369–370, 372 passim
Moores Cancer Center, University of California at San Diego (2015) Microscopy – protocols. Moores Cancer Center, University of California at San Diego, San Diego, CA, http://healthsciences.ucsd.edu/research/moores/shared-resources/microscopy/Pages/protocols.aspx. Accessed on 10 Oct 2015
Acknowledgments
R. S. H. is supported by the Stowers Institute for Medical Research and is an American Cancer Research Professor supported by the award RP-05-086-06DDC. Original data underlying this manuscript can be accessed from the Stowers Original Data Repository at http://www.stowers.org/research/publications/libpb-1019.
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Hughes, S.E., Hawley, R.S. (2017). Live Imaging of Meiosis I in Late-Stage Drosophila melanogaster Oocytes. In: Stuart, D. (eds) Meiosis. Methods in Molecular Biology, vol 1471. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6340-9_14
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DOI: https://doi.org/10.1007/978-1-4939-6340-9_14
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