Summary
Appropriate culture of murine embryonic stem cells is critical to enabling introduced mutations to be passed through the germ line. ES cells must be carefully cultured to ensure that pluripotency is maintained. The feeder cells and foetal calf serum used to culture the cells can significantly influence germ line transmission potential. Additionally, ES cells can be karyotypically unstable, so determining the karyotype of ES cell lines will increase your confidence in the ability of the manipulated cells to contribute to the germ line in chimaeric mice.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Martin, G.R. (1981). Isolation of a pluripotent cell line from early mouse embryos cultured in medium conditioned by teratocarcinoma stem cells. Proc. Natl. Acad. Sci. U.S.A. 78, 7634–7638
Evans, M.J. and Kaufman, M.H. (1981). Establishment in culture of pluripotential cells from mouse embryos. Nature 292, 154–156
Williams, R.L., Hilton, D.J., Pease, S., Willson, T.A, Stewart, C.L., Gearing, D.P., Wagner, E.F., Metcalf, D., Nicola, N.A., and Gough, N.M. (1988). Myeloid leukaemia inhibitory factor maintains the developmental potential of embryonic stem cells. Nature 336, 684–687
Smith, A.G., Heath, J.K., Donaldson, D.D., Wong, G.G., Moreau, J., Stahl, M., and Rogers, D. (1988). Inhibition of pluripotential embryonic stem cell differentiation by purified polypeptides. Nature 336, 688–690
Solter, D. (2006). From teratocarcinomas to embryonic stem cells and beyond: a history of embryonic stem cell research. Nat. Rev. Genet. 7, 319–327
Nagy, A. (2008). URL: www.mshri.on.ca/nagy/r1.htm retrieved on 28th February 2008
Wellcome Trust Sanger Institute. (2008). URL: www.sanger.ac.uk/Software/archives/mouse-help.shtml retrieved on 28th February 2008
Keskintepe, L., Norris, K., Pacholczyk, G., Dederscheck, S.M., and Eroglu, A. (2007). Derivation and comparison of C57BL/6 embryonic stem cells to a widely used 129 embryonic stem cell line. Transgenic Res. 16, 751–758
Blake, M.S., Johnston, K.H., Russell-Jones, G.J., and Gotschlich, E.C. (1984). A rapid, sensitive method for detection of alkaline phosphatase-conjugated anti-antibody on Western blots. Anal. Biochem. 136, 175–179
Ayral, A., Clarkson, S., Cheeseman, M., Wells, S., and Dear, T. (2006). A panel of optimized primers and positive control DNAs for PCR detection of common biological contaminants in mouse cell lines and tissue samples. Lab Anim. 35, 31–36
Udy, G.B., Parkes, B.D., and Wells, D.N. (1997). ES cell cycle rates affect gene targeting frequencies. Exp. Cell Res.231, 296–301
Balbach, S.T., Jauch, A., Böhm-Steuer, B., Cavaleri, F.M., Han, Y.M., and Boiani M. (2007). Chromosome stability differs in cloned mouse embryos and derivative ES cells. Dev. Biol. 308, 309–321
Longo, L., Bygrave, A., Grosveld, F.G., and Pandolfi, P.P. (1997). The chromosome make-up of mouse embryonic stem cells is predictive of somatic and germ cell chimaerism. Transgenic Res. 6, 321–328
Liu, X., Wu, H., Loring, J., Hormuzdi, S., Disteche, C.M., Bornstein, P., and Jaenisch R. (1997). Trisomy eight in ES cells is a common potential problem in gene targeting and interferes with germ line transmission. Dev. Dyn. 209, 85–91
Hook, E.B. (1977). Exclusion of chromosomal mosaicism: Tables of 90%, 95%, and 99% confidence limits and comments on use. Am. J. Hum. Genet. 29, 94–97
Nicklas, W., and Weiss. J. (2000). Survey of embryonic stem cells for murine infective agents. Comp. Med. 50, 410–411
Kyuwa, S. (1997). Replication of murine coronaviruses in mouse embryonic stem cell lines in vitro. Exp. Anim. 46, 311–313
Lindsey, J., Boorman, G., Collins, M., Hsu, C.-K., Van Hoosier., Jr.,G., and Wagner, J. (1991) Infectious Diseases in Mice and Rats. National Academy Press, Washington. DC. pp. 42–48
Tucker, K.L., Wang, Y., Dausman, J., and Jaenisch, R. (1997). A transgenic mouse strain expressing four drug-selectable marker genes. Nucleic Acids Res.25, 3745–3746
O’Gorman, S., Dagenais, N.A., Qian, M., and Marchuk, Y. (1997). Protamine-Cre recombinase transgenes efficiently recombine target sequences in the male germ line of mice, but not in embryonic stem cells. Proc. Natl. Acad. Sci. U.S.A. 94, 14602–14607
Acknowledgements
We thank Monica Neilan for assistance with ES cell culture and Adam Steinberg and the Biochemistry Medialab of the University of Wisconsin for providing the mouse image.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2009 Humana Press, a part of Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Barbaric, I., Dear, T.N. (2009). Culture of Murine Embryonic Stem Cells. In: Cartwright, E. (eds) Transgenesis Techniques. Methods in Molecular Biology, vol 561. Humana Press. https://doi.org/10.1007/978-1-60327-019-9_11
Download citation
DOI: https://doi.org/10.1007/978-1-60327-019-9_11
Published:
Publisher Name: Humana Press
Print ISBN: 978-1-60327-018-2
Online ISBN: 978-1-60327-019-9
eBook Packages: Springer Protocols