Lineage analysis is widely used because it provides a very powerful tool for characterizing the developmental behavior of the cells in vivo. In this chapter, we describe a particularly informative variant of lineage analysis that we term “single-cell lineage analysis”. As in traditional lineage analysis, the method employs a Tamoxifen (Tmx)-inducible CAGCreER mouse line, which is crossed to an R26R reporter line that can be activated by Cre-mediated DNA recombination. However, instead of driving CreER at a high level within a subset of cells defined by a particular promoter, CreER is driven with a generic promoter that is active in essentially all cells throughout the lifespan of the mouse. Specificity comes from using only a very low dose of Tmx so that just a few random, widely separated individual cells undergo recombination and become labeled. The growth and behavior of most such initially marked cells can subsequently be followed over time because each one forms a growing clone of marked cells that does not overlap with other clones due to their rarity. Following individual cell growth patterns provides much more information than can be derived from traditional lineage analysis, which relies on promoter specificity and uses high doses of Tmx that cannot resolve the behavior of single cells. We illustrate the value of single-cell lineage analysis using a recent study of fetal germ cell development and a recent search for female germ-line stem cells in adult mouse ovaries.
Feil S, Valtcheva N, Feil R (2009) Inducible Cre mice. Gene Knockout Protoc 530:343–363.CrossRefGoogle Scholar
Vasioukhin V, Degenstein L, Wise B et al (1999) The magical touch: genome targeting in epidermal stem cells induced by tamoxifen application to mouse skin. Proc Natl Acad Sci U S A 96:8551–8556.CrossRefPubMedPubMedCentralGoogle Scholar
Harfe BD, Scherz PJ, Nissim S et al (2004) Evidence for an expansion-based temporal Shh gradient in specifying vertebrate digit identifies. Cell 118:517–528.CrossRefPubMedGoogle Scholar
Barker N, van Es JH, Kuipers J et al (2007) Identification of stem cells in small intestine and colon by marker gene Lgr5. Nature 449:1003–1007.CrossRefPubMedGoogle Scholar
Nakagawa T, Nabeshima Y, Yoshida S et al (2007) Functional identification of the actual and potential stem cell compartments in mouse spermatogenesis. Dev Cell 12:195–206.CrossRefPubMedGoogle Scholar
Zhou BO, Yue R, Murphy MM et al (2014) Leptin-receptor-expressing mesenchymal stromal cells represent the main source of bone formed by adult bone marrow. Cell Stem Cell 15:154–168.CrossRefPubMedPubMedCentralGoogle Scholar
Hayashi S, McMahon AP (2002) Efficient recombination in diverse tissues by a tamoxifen-inducible form of Cre: a tool for temporally regulated gene activation/inactivation in the mouse. Dev Biol 244:305–318.CrossRefPubMedGoogle Scholar