In Vivo Genetic Alteration and Lineage Tracing of Single Stem Cells by Live Imaging. Farrelly, O., Kuri, P., & Rompolas, P. Methods in molecular biology (Clifton, N.J.), 1879:1–14, 2019. doi abstract bibtex Studies characterizing stem cell lineages in different organs aim to understand which cells particular progenitors can give rise to and how this process is controlled. Because the skin contains several resident stem cell populations and undergoes constant turnover, it is an ideal tissue in which to study this phenomenon. Furthermore, with the advent of two-photon microscopy techniques in combination with genetic tools for cell labeling, this question can be studied non-invasively by using live imaging. In this chapter, we describe an experimental approach that takes this technique one step further. We combine the Cre and Tet inducible genetic systems for single clone labeling and genetic manipulation in a specific stem cell population in the skin by using known drivers. Our system involves the use of gain- and loss-of-function alleles activated only in a differentially labeled population to distinguish single clones. The same region within a tissue is imaged repeatedly to document the fate and interactions of single clones with and without genetic modifications in the long term. Implementing this lineage tracing approach while documenting changes in cell behavior brought about by the genetic alterations allows both aspects to be linked. Because of the inherent flexibility of the approach, we expect it to have broad applications in studying stem cell function not only in the skin, but also in other tissues amenable to live imaging.
@article{farrelly_vivo_2019,
title = {In {Vivo} {Genetic} {Alteration} and {Lineage} {Tracing} of {Single} {Stem} {Cells} by {Live} {Imaging}},
volume = {1879},
doi = {10.1007/7651_2018_172 [doi]},
abstract = {Studies characterizing stem cell lineages in different organs aim to understand which cells particular progenitors can give rise to and how this process is controlled. Because the skin contains several resident stem cell populations and undergoes constant turnover, it is an ideal tissue in which to study this phenomenon. Furthermore, with the advent of two-photon microscopy techniques in combination with genetic tools for cell labeling, this question can be studied non-invasively by using live imaging. In this chapter, we describe an experimental approach that takes this technique one step further. We combine the Cre and Tet inducible genetic systems for single clone labeling and genetic manipulation in a specific stem cell population in the skin by using known drivers. Our system involves the use of gain- and loss-of-function alleles activated only in a differentially labeled population to distinguish single clones. The same region within a tissue is imaged repeatedly to document the fate and interactions of single clones with and without genetic modifications in the long term. Implementing this lineage tracing approach while documenting changes in cell behavior brought about by the genetic alterations allows both aspects to be linked. Because of the inherent flexibility of the approach, we expect it to have broad applications in studying stem cell function not only in the skin, but also in other tissues amenable to live imaging.},
language = {eng},
journal = {Methods in molecular biology (Clifton, N.J.)},
author = {Farrelly, O. and Kuri, P. and Rompolas, P.},
year = {2019},
pmid = {30242568. NIHMSID: 1612800. CPAT Core (A), STaR Core (B), Mentoring Sub-Core.},
keywords = {Admin Core, CPAT Core, Mentoring Sub-Core, STaR Core},
pages = {1--14},
}
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