Efficient derivation of stable primed pluripotent embryonic stem cells from bovine blastocysts

TLDR

Embryonic stem cells are derived from the inner cell mass of blastocysts, and stable bovine ESC lines are sought for genomic testing, selection, genome engineering, and disease modeling, yet producing them remains a challenge. Using a culture medium containing fibroblast growth factor 2 and a canonical Wnt‑signaling inhibitor, the authors established bovine ESCs with stable morphology, transcriptome, karyotype, proliferation, pluripotency markers, and epigenetic features. The protocol yielded 100 % efficient, rapidly established (3–4 weeks) bESC lines that are easy to propagate and, when used as nuclear transfer donors, produced normal blastocysts, enabling genomic selection, genome editing, and high‑value cattle production.

Abstract

Embryonic stem cells (ESCs) are derived from the inner cell mass of preimplantation blastocysts. From agricultural and biomedical perspectives, the derivation of stable ESCs from domestic ungulates is important for genomic testing and selection, genome engineering, and modeling human diseases. Cattle are one of the most important domestic ungulates that are commonly used for food and bioreactors. To date, however, it remains a challenge to produce stable pluripotent bovine ESC lines. Employing a culture system containing fibroblast growth factor 2 and an inhibitor of the canonical Wnt-signaling pathway, we derived pluripotent bovine ESCs (bESCs) with stable morphology, transcriptome, karyotype, population-doubling time, pluripotency marker gene expression, and epigenetic features. Under this condition bESC lines were efficiently derived (100% in optimal conditions), were established quickly (3-4 wk), and were simple to propagate (by trypsin treatment). When used as donors for nuclear transfer, bESCs produced normal blastocyst rates, thereby opening the possibility for genomic selection, genome editing, and production of cattle with high genetic value.