Additional file 1.
Identification and validation of inducible/exchangeable recombinant mES clones. (a) Recombinant mES clones were identified by PCR analysis. (b) q-PCR analysis and Luciferase assays using Dual Luciferase Reporter Assay System was performed on mES clones overexpressing the firefly luciferase (Luc) gene. The system was activated upon the removal of Tc (after 17, 24, 39 and 48 hours) from the medium. Protein extracts of mES cells were prepared at the same time points and luminescence quantified. (c) q-PCR analysis and YFP fluorescence assay to detect the expression of the YFP reporter. (d) Expression of mES cells overexpressing Luc after 24 hours from the complete removal of Tc from the medium; the degree of induction was easily manipulated by titrating the Tc. (e) Expression profile (q-PCR) of the pluripotency gene Oct3/4, and of markers of the mesoderm (Brachyury), ectoderm (Gfap) and endoderm (Afp) during differentiation of EB3 and of the parent cell line (E14).
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De Cegli et al. Genome Biology 2010 11:R64 doi:10.1186/gb-2010-11-6-r64