Characterization of bovine Nanog 5'-flanking region during differentiation of mouse embryonic stem cells

초록

Embryonic stem cells (ESCs) have been used as a powerful tool for use in research involving gene manipulated animal models and the study of developmental gene regulation. Among the critical regulatory factors in undifferentiated embryonic stem cells maintaining the pluripotency and self-renewal, the Nanog gene may play a very important role for maintenance of the pluripotency and self-renewal. Nevertheless, the studies of the Nanog gene from domestic animals including bovine for derivation of embryonic stem cells have been difficult since pluripotency maintaining factors and specific markers for live stock ESCs have not been probed yet. Therefore, in this study, in order to understand the function and mechanisms of regulation of the livestock Nanog gene using validated mouse ESCs, we cloned 600bp (-420/+181) bovine Nanog 5’-flaking region, and tagged it with humanized renilla reniformis green fluorescence protein (hrGFP) as a tracing reporter. After the construct was integrated into mouse genomic DNA of ESCs, GFP expression for bovine Nanog promoter was very high in the mouse ESC line. GFP expression is monitored upon ESC differentiation and gradually reduced along with differentiation toward neuron and adipocyte using retinoic acid. Activity of bovine nanog (-420/+181) promoter was compared with already known mouse and human nanog promoters in mouse ESC and their regulations were likely to have a very similar pattern. In conclusion, bovine nanog 5'-flanking region is working in mouse ES cells and has similar characteristic as those of mouse and human. These results suggest that bovine gene function studied in mouse ES cells is chased and extrapolated which is applicate into bovine ES cells chacterization.