SRA

Despite four decades of effort, robust propagation of pluripotent stem cells from livestock animals remains challenging. The requirements for self-renewal are unclear and the relationship of cultured stem cells to pluripotent cells resident in the embryo uncertain. Here we avoided feeder cells or serum factors to provide a defined culture microenvironment. We show that the combination of Activin A, Fibroblast growth factor, and Wnt inhibitor XAV939 (AFX), supports establishment and continuous expansion of pluripotent stem cell lines from porcine, ovine and bovine embryos. Germ layer differentiation was evident in teratomas and readily induced in vitro. Global transcriptome analyses highlighted commonality in transcription factor expression across the three species, while global comparison with porcine embryo stages showed proximity to bilaminar disc epiblast. Clonal genetic manipulation and gene targeting were exemplified in porcine stem cells. We further demonstrated that genetically modified AFX stem cells gave rise to cloned porcine foetuses by nuclear transfer. In summary, for major livestock mammals pluripotent stem cells related to the formative embryonic disc are reliably established using a common and defined signaling environment. Overall design: RNA-seq analysis of EDSCs established from pig, sheep and cow embryos