Summary
Complementing mutant embryos or embryonic stem cells with normal cells in embryonic chimeras is a valuable tool for investigating phenotypes. Chimera approaches provide a method to examine the phenotype of mutant cells, including hematopoiesis, in mutants with early embryonic lethality. Complementation with normal cells in a chimera can, in most instances, rescue mutant cells to later stages of gestation and beyond, permitting analysis of contribution and function of mutant cells in various organs, both within the chimera, but also by using functional transplantation assays for hematopoietic stem and progenitor cells. This chapter describes principles and methods for the generation of mouse chimeras, for identification and quantitative analysis of cell contribution in chimeras, and for chimeric fetal liver transplantation into adult recipients and analysis of mutant cells in the adult.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Feil R. (2007) Conditional somatic mutagenesis in the mouse using site- specific recombinases. Handb Exp Pharmacol 178, 3–28.
Matsumoto N., Kubo A., Liu h., Akita K., Laub F., Ramirez F., Keller G., Friedman S. L. (2006) Developmental regulation of yolk sac hematopoiesis by Kruppel- like factor 6. Blood 107, 1357–1365.
Tam P. P., Rossant J. (2003) Mouse embryonic chimeras: tools for studying mammalian development. Development 130, 6155–6163.
Chen J., Lansford R., Stewart V., Young F., Alt F. W. (1993) RAG- 2- deficient blastocyst complementation: an assay of gene function in lymphocyte development. Proc Natl Acad Sci USA 90, 4528–4532.
Wang L. C., Swat W., Fujiwara Y., Davidson L., Visvader J., Kuo F., Alt F. W., Gilliland D. G., Golub T. R., Orkin S. h. (1998) The TEL/ETV6 gene is required specifically for hematopoiesis in the bone marrow. Genes Dev 12, 2392–2402.
Ernst P., Fisher J. K., Avery W., Wade S., Foy D., Korsmeyer S. J. (2004) Definitive hematopoiesis requires the mixed- lineage leukemia gene. Dev Cell 6, 437–443.
Tsang A. P., Fujiwara Y., Hom D. B., Orkin S. h. (1998) Failure of megakaryopoiesis and arrested erythropoiesis in mice lacking the GATA- 1 transcriptional cofactor FOG. Genes Dev 12, 1176–1188.
Eckardt S., Leu N. A., Bradley h. L., Kato h., Bunting K. D., Mclaughlin K. J. (2007) Hematopoietic reconstitution with androgenetic and gynogenetic stem cells. Genes Dev 21, 409–419.
Nagy A., Gertsenstein M., Vintersten K., Behringer R. (2003) Manipulating the Mouse Embryo, 3rd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.
Abbondanzo S. J., Gadi I., Stewart C. L. (1993) Derivation of embryonic stem cell lines, in Methods Enzymol 225: Guide to Techniques in Mouse Development (Wassarman, P.M, DePamphilis, M.L., eds.), Academic Press, San Diego, CA, pp. 803–823.
Okabe M., Ikawa M., Kominami K., Nakanishi T., Nishimune Y. (1997) ‘Green mice’ as a source of ubiquitous green cells. FEBS Lett 407, 313–319.
Kawakami N., Sakane N., Nishizawa F., Iwao M., Fukada S. I., Tsujikawa K., Kohama Y., Ikawa M., Okabe M., Yamamoto h. (1999) Green fluorescent protein- transgenic mice: immune functions and their application to studies of lymphocyte development. Immunol Lett 70, 165–171.
Spangrude G. J., Cho S., Guedelhoefer O., Vanwoerkom R. C., Fleming W. h. (2006) Mouse models of hematopoietic engraftment: limitations of transgenic green fluorescent protein strains and a high- performance liquid chromatography approach to analysis of erythroid chimerism. Stem Cells 24, 2045–2051.
Wang Z., Jaenisch R. (2004) At most three ES cells contribute to the somatic lineages of chimeric mice and of mice produced by ES- tetraploid complementation. Dev Biol 275, 192–201.
Yagi T., Tokunaga T., Furuta Y., Nada S., Yoshida M., Tsukada T., Saga Y., Takeda N., Ikawa Y., Aizawa S. (1993) A novel ES cell line, TT2, with high germline- differentiating potency. Anal Biochem 214, 70–76.
Stewart C. L. (1993) Production of chimeras between embryonic stem cells and embryos, in Methods Enzymol 225: Guide to Techniques in Mouse Development (Wassarman, P.M, DePamphilis, M.L., eds.), Academic Press, San Diego, CA, pp. 823–855.
Mann J. R. (1993) Surgical Techniques in Production of Transgenic Mice, in Methods Enzymol 225: Guide to Techniques in Mouse Development (Wassarman, P.M, DePamphilis, M.L., eds.), Academic Press, San Diego, CA, pp. 782–793.
Nagy A., Rossant J. (1993) Production of completely ES cell derived fetuses, in Gene Targeting, 1 ed. IRL (Joyner A.L., ed.), Oxford, pp. 147–178.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2008 Humana Press, a part of Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Eckardt, S., McLaughlin, K.J. (2008). Transplantation of Chimeric Fetal Liver to Study Hematopoiesis. In: Bunting, K.D. (eds) Hematopoietic Stem Cell Protocols. Methods in Molecular Biology™, vol 430. Humana Press. https://doi.org/10.1007/978-1-59745-182-6_14
Download citation
DOI: https://doi.org/10.1007/978-1-59745-182-6_14
Publisher Name: Humana Press
Print ISBN: 978-1-58829-868-3
Online ISBN: 978-1-59745-182-6
eBook Packages: Springer Protocols