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Generation of induced pluripotent stem cells from domestic goats
Author(s) -
Sandmaier Shelley E. S.,
Nandal Anjali,
Powell Anne,
Garrett Wesley,
Blomberg Leann,
Donovan David M.,
Talbot Neil,
Telugu Bhanu P.
Publication year - 2015
Publication title -
molecular reproduction and development
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.745
H-Index - 105
eISSN - 1098-2795
pISSN - 1040-452X
DOI - 10.1002/mrd.22512
Subject(s) - biology , sox2 , induced pluripotent stem cell , reprogramming , klf4 , homeobox protein nanog , embryonic stem cell , stem cell , microbiology and biotechnology , leukemia inhibitory factor , kosr , viral vector , genetics , cell , gene , recombinant dna
SUMMARY The creation of genetically modified goats provides a powerful approach for improving animal health, enhancing production traits, animal pharming, and for ensuring food safety all of which are high‐priority goals for animal agriculture. The availability of goat embryonic stem cells (ESCs) that are characteristically immortal in culture would be of enormous benefit for developing genetically modified animals. As an alternative to long‐sought goat ESCs, we generated induced pluripotent stem cells (iPSC) by forced expression of bovine POU5F1, SOX2, MYC, KLF4, LIN‐28 , and NANOG reprogramming factors in combination with a MIR302/367 cluster, delivered by lentiviral vectors. In order to minimize integrations, the reprogramming factor coding sequences were assembled with porcine teschovirus‐1 2A (P2A) self‐cleaving peptides that allowed for tri‐cistronic expression from each vector. The lentiviral‐transduced cells were cultured on irradiated mouse feeder cells in a semi‐defined, serum‐free medium containing fibroblast growth factor (FGF) and/or leukemia inhibitory factor (LIF). The resulting goat iPSC exhibit cell and colony morphology typical of human and mouse ESCs—that is, well‐defined borders, a high nuclear‐to‐cytoplasmic ratio, a short cell‐cycle interval, alkaline phosphatase expression, and the ability to generate teratomas in vivo. Additionally, these goat iPSC demonstrated the ability to differentiate into directed lineages in vitro. These results constitute the first steps in establishing integration and footprint‐free iPSC from ruminants. Mol. Reprod. Dev. 82: 709–721, 2015. © 2015 Wiley Periodicals, Inc .