Figure 1.

Attempts to Obtain Chicken Reprogrammed Cells with Various Delivery Systems

Primary chicken embryonic fibroblasts (CEFs) were infected with the pLent polycistronic lentivirus. Colonies emerged (A) and expressed GFP (B), indicating that they were infected, although they failed to be amplified. More than 50 colonies were isolated in several independent experiments. Similarly, CEFs were infected with a cocktail of individual lentiviruses carrying mouse Oct4, Sox2, Klf4, and c-Myc cDNAs. Compact colonies with AP-positive cells were obtained (C and D) but could not be established long term. In another attempt, CEFs were infected with pMX retroviruses carrying mouse Oct4, Sox2, Klf4, and c-Myc cDNAs. The titer was determined using pMX-GFP as shown in Figure S1. Morphological changes were rapidly observed (E), and colonies expanded once individually picked (F). The phenotypic changes were not stable, and cells reverted to a fibroblast phenotype (G) after four to six passages. Similarly, CEFs infected with Sendai viruses carrying human OCT4, SOX2, KLF4, and c-MYC cDNAs exhibited morphological changes (H) but failed to be established and reverted to a fibroblast phenotype after a few passages.

Scale bars, 50 μm.