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Introduction of cell markers into germ layer tissues of the mouse gastrula by whole embryo electroporation
Author(s) -
Davidson Bruce P.,
Tsang Tania E.,
Khoo PohLynn,
Gad Jacqueline M.,
Tam Patrick P.L.
Publication year - 2003
Publication title -
genesis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.093
H-Index - 110
eISSN - 1526-968X
pISSN - 1526-954X
DOI - 10.1002/gene.10166
Subject(s) - electroporation , gastrulation , epiblast , biology , germ layer , microbiology and biotechnology , embryo , endoderm , reporter gene , green fluorescent protein , nodal signaling , cellular differentiation , genetics , embryonic stem cell , gene expression , gene , embryogenesis , induced pluripotent stem cell
Summary: We have optimized the technique of electroporation for introducing genetic markers into cells of the gastrulating mouse embryo to follow cell fates, tissue movement, and lineage differentiation. Using a plate‐needle electrode combination and specific route of plasmid delivery, labeling could be targeted to discrete regions of the epiblast or the endoderm of the late gastrula. Among the various types of fluorescent and chromogenic reporter constructs tested, those driven by CMV promoter are efficient and strong expression can be detected as soon as 2–3 h after electroporation. The efficacy of marking cell lineages by CRE‐mediated activation of reporters proved to be inefficient for tracking cell lineages due to an obligatory 8–9‐h lag from the electroporation of constructs to the expression of reporter. This significant time lag also raises concern of the temporal precision at which tissue‐ or stage‐specific knock‐out or activation of genetic activity may be achieved by the Cre‐loxP mechanism. genesis 35:57–62, 2003. © 2002 Wiley‐Liss, Inc.