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. Author manuscript; available in PMC: 2014 Jun 16.
Published in final edited form as: Stem Cells. 2008 Jun 5;26(8):2131–2141. doi: 10.1634/stemcells.2008-0388

Figure 1.

Figure 1

Cre-loxP-based, enhanced green fluorescent protein-inducible assay for reprogramming (CLEAR). (A): A diagrammatic illustration of CLEAR analysis. CIPOE NSC lines were established by infection of adult NSCs derived from transgenic mice harboring Oct4-enhanced green fluorescent protein reporter with retroviruses to coexpress the Cre recombinase and puromycin resistance gene. Z-Red ESCs carry an inducible DsRed expression cassette upon Cre-mediated excision. PEG-induced fusion of Z-Red ESCs and CIPOE NSCs leads to GFP expression as an indicator of Oct4 reactivation and DsRed expression as a reporter for fusion events. The dual-color reporter system can be monitored by both live fluorescence microscopy and quantitative flow cytometry to probe reprogramming processes. (B, C): Live images of fused ES-like colonies. Shown are sample images of fused ES-like colonies at 48 hours (B) and 96 hours (C) after PEG-induced fusion between CIPOE NSCs and Z-Red ESCs. Arrows point to DsRed+GFP− cells that were successfully fused but incompletely reprogrammed. Scale bar = 20 μm. Abbreviations: ES, embryonic stem; GFP, green fluorescent protein; ires, internal ribosome entry site; NSC, neural stem cell/progenitor; O4E, Oct4-enhanced green fluorescent protein; PEG, polyethylene glycol.