Abstract
The mouse (Mus musculus) represents the central mammalian genetic model system for biomedical and developmental research. Mutant mouse models have provided important insights into chromosome dynamics during the complex meiotic differentiation program that compensates for the genome doubling at fertilization. Homologous chromosomes (homologues) undergo dynamic pairing and recombine during first meiotic prophase before they become partitioned into four haploid sets by two consecutive meiotic divisions that lack an intervening S-phase. Fluorescence in situ hybridization (FISH) has been instrumental in the visualization and imaging of the dynamic reshaping of chromosome territories and mobility during prophase I, in which meiotic telomeres were found to act as pacemakers for the chromosome pairing dance. FISH combined with immunofluorescence (IF) co-staining of nuclear proteins has been instrumental for the visualization and imaging of mammalian meiotic chromosome behavior. This chapter describes FISH and IF methods for the analysis of chromosome dynamics in nuclei of paraffin-embedded mouse testes. The techniques have proven useful for fresh and archived paraffin testis material of several mammalian species.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Rasmussen SW, Holm PB (1980) Mechanics of meiosis. Hereditas 93(2):187–216
Scherthan H (2006) Meiotic telomeres. In: de Lange T, Zakian V, Blackburn E (eds) Telomeres, 2nd edn. CSH Press, Cold Spring Harbor, NY, pp 225–259
Gerton JL, Hawley RS (2005) Homologous chromosome interactions in meiosis: diversity amidst conservation. Nat Rev Genet 6(6):477–487
Ishiguro K et al (2014) Meiosis-specific cohesin mediates homolog recognition in mouse spermatocytes. Genes Dev 28(6):594–607
Boateng KA, Bellani MA, Gregoretti IV, Pratto F, Camerini-Otero RD (2013) Homologous pairing preceding SPO11-mediated double-strand breaks in mice. Dev Cell 24(2):196–205
Scherthan H et al (1996) Centromere and telomere movements during early meiotic prophase of mouse and man are associated with the onset of chromosome pairing. J Cell Biol 134(5):1109–1125
Liebe B et al (2006) Mutations that affect meiosis in male mice influence the dynamics of the mid-preleptotene and bouquet stages. Exp Cell Res 312(19):3768–3781
Shibuya H, Morimoto A, Watanabe Y (2014) The dissection of meiotic chromosome movement in mice using an in vivo electroporation technique. PLoS Genet 10(12), e1004821
Scherthan H, Schofisch K, Dell T, Illner D (2014) Contrasting behavior of heterochromatic and euchromatic chromosome portions and pericentric genome separation in pre-bouquet spermatocytes of hybrid mice. Chromosoma 123(6):609–624
Ahmed EA, Sfeir A, Takai H, Scherthan H (2013) Ku70 and non-homologous end joining protect testicular cells from DNA damage. J Cell Sci 126(Pt 14):3095–3104
Bordlein A et al (2011) SPOC1 (PHF13) is required for spermatogonial stem cell differentiation and sustained spermatogenesis. J Cell Sci 124(Pt 18):3137–3148
Novak I et al (2008) Cohesin Smc1beta determines meiotic chromatin axis loop organization. J Cell Biol 180(1):83–90
Lichter P, Cremer T (1992) Chromosome analysis by non-isotopic in situ hybridization. In: Rooney DE, Czepulkowsky BH (eds) Human cytogenetics – a practical approach, 2nd edn. IRL Press, Oxford
Roche PJ (2006) Preparation of template DNA and labeling techniques. Methods Mol Biol 326:9–16
Scherthan H, Cremer T (1994) Methodology of non-isotopic in situ-hybridization in paraffin embedded tissue sections. In: Adolph KW (ed) Methods in molecular genetics, vol 5, Academic. San Diego, CA, pp 223–238
Schmitt E, Hausmann M (2010) COMBO FISH. Methods Mol Biol 659:185–202
Lansdorp PM (1996) Close encounters of the PNA kind. Nat Biotechnol 14(13):1653
Florijn RJ, Slats J, Tanke HJ, Raap AK (1995) Analysis of antifading reagents for fluorescence microscopy. Cytometry 19:177–182
Burck H-C (1988) Histologische Technik [Histological technique]. Thieme, Stuttgart
Hopman AH, van Hooren E, van de Kaa CA, Vooijs PG, Ramaekers FC (1991) Detection of numerical chromosome aberrations using in situ hybridization in paraffin sections of routinely processed bladder cancers. Mod Pathol 4(4):503–513
Morelli MA, Werling U, Edelmann W, Roberson MS, Cohen PE (2008) Analysis of meiotic prophase I in live mouse spermatocytes. Chromosome Res 16(5):743–760
Shi SR, Key ME, Kalra KL (1991) Antigen retrieval in formalin-fixed, paraffin-embedded tissues: an enhancement method for immunohistochemical staining based on microwave oven heating of tissue sections. J Histochem Cytochem 39(6):741–748
Scherthan H et al (1994) Comparative chromosome painting discloses homologous segments in distantly related mammals. Nat Genet 6(4):342–347
Meinkoth J, Wahl G (1984) Hybridization of nucleic acids immobilized on solid supports. Anal Biochem 138(2):267–284
Scherthan H et al (2000) Meiotic telomere distribution and sertoli cell nuclear architecture are altered in Atm- and Atm-p53-deficient mice. Mol Cell Biol 20(20):7773–7783
Acknowledgments
This work was supported by a grant from the Deutsche Forschungsgemeinschaft (SCHE350/10-1, SPP 1384). I thank Doris Illner for critical reading of the manuscript.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2017 Springer Science+Business Media New York
About this protocol
Cite this protocol
Scherthan, H. (2017). Imaging of Chromosome Dynamics in Mouse Testis Tissue by Immuno-FISH. 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_12
Download citation
DOI: https://doi.org/10.1007/978-1-4939-6340-9_12
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-6338-6
Online ISBN: 978-1-4939-6340-9
eBook Packages: Springer Protocols