Derivation and characterization of bovine embryonic stem cells in a commercially available culture condition

NAVARRO M; ROSS PJ; MUTTO AA

Buenos Aires

Congreso; Tenth Congress of the Latin American Society for Developmental Biology; 2020

Latin American Society for Developmental Biology

Derivation of bovine embryonic stem cells (bESCs) from blastocyst stage embryos was recently reported. bESCs were established in a custom-made mTeSR medium devoid of TGFß supplemented with FGF2 and IWR1, a WNT signaling inhibitor. However, this medium is not commercially available which makes it inaccessible to most laboratories. Thus, the aim of this work was to derive bESC in a commercially available base medium, adapting the conditions described previously. Single zona-free IVF blastocysts were plated over an irradiated mouse feeder layer (MEF) and cultured in the N2B27 culture medium, supplemented with FGF2 and IWR1. The whole culture was passaged once a week onto a fresh MEF by single cell dissociation. Once established, cells were also adapted to feeder-free conditions (FF).After 4 weeks, well defined colonies appeared with an efficiency of 30%. bESC cultured over MEF as well as FF, were alkaline phosphatase positive, expressed OCT4 and SOX2 markers, differentiated in vitro into the three germ layers, and presented a normal chromosome number. Additionally, cells retained stemness and genome stability in long term-culture (>35passages).These results suggest that bESC can be derived and stably maintained is this commercially available medium without losing their pluripotency and proliferation potential. bESC can now be produced in a simplified culture condition, representing an important advance for genomic testing and selection, genome engineering, and for studying human diseases.