Developmental analyses of mouse embryos and adults using a non-overlapping tracing system for all three germ layers

Takashi Serizawa, Ayako Isotani, Takafumi Matsumura, Katsuyuki Nakanishi, Shigenori Nonaka, Shinsuke Shibata, Masahito Ikawa, Hideyuki Okano

Research output: Contribution to journalArticle

Abstract

Genetic lineage-tracing techniques are powerful tools for studying specific cell populations in development and pathogenesis. Previous techniques have mainly involved systems for tracing a single gene, which are limited in their ability to facilitate direct comparisons of the contributions of different cell lineages. We have developed a new combinatorial system for tracing all three germ layers using self-cleaving 2A peptides and multiple site-specific recombinases (SSRs). In the resulting TRiCK (TRiple Coloured germ layer Knock-in) mice, the three germ layers are conditionally and simultaneously labelled with distinct fluorescent proteins via embryogenesis. We show that previously reported ectopic expressions of lineage markers are the outcome of secondary gene expression. The results presented here also indicate that the commitment of caudal axial stem cells to neural or mesodermal fate proceeds without lineage fluctuations, contrary to the notion of their bi-potency. Moreover, we developed IMES, an optimized tissue clearing method that is highly compatible with a variety of fluorescent proteins and immunostaining, and the combined use of TRiCK mice and IMES can facilitate comprehensive analyses of dynamic contributions of all three germ layers.

Original languageEnglish
Article numberdev174938
JournalDevelopment (Cambridge)
Volume146
Issue number21
DOIs
Publication statusPublished - 2019

Keywords

  • 2A peptide
  • Fate map
  • Germ layer
  • Lineage-tracing system
  • Site-specific recombinase
  • Tissue clearing

ASJC Scopus subject areas

  • Molecular Biology
  • Developmental Biology

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