期刊
DEVELOPMENT
卷 146, 期 21, 页码 -出版社
COMPANY BIOLOGISTS LTD
DOI: 10.1242/dev.174938
关键词
Lineage-tracing system; Fate map; Germ layer; 2A peptide; Site-specific recombinase; Tissue clearing
资金
- Keio University
- Osaka University
- National Institute for Basic Biology
- Program for the Advancement of Research in Core Projects on Longevity of the Keio University Global Research Institute from Keio University
- Japan Society for the Promotion of Science KAKENHI Grant [JP16H06280]
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.
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