Isolation and Characterization of Cultured Chicken Oviduct Epithelial Cells and In Vitro Validation of Constructed Ovalbumin Promoter in these Cells

Animal Biotechnology Division, National Institute of Animal Science, Rural Development Administration, 1500, Kongjwipatjwi-ro, Iseo-myeon, Wanju-gun, Jeollabuk-do, 55365, Republic of Korea

Correspondence: Sung June Byun, Tel: +82-63-238-7263, Fax: +82-63-238-7297, Email: pcs1778@korea.ac.kr

Received: 4 September 2020 • Revised: 25 November 2020 • Accepted: 7 December 2020

Abstract

Objective
Transgenic hens hold a great promise for the production of various valuable protein. Through virus transduction into stage X embryo, the transgene expression under the control of constructed chicken ovalbumin promoters has been successfully achieved. However, absence of the validation system that can evaluate differently developed ovalbumin promoters in in vitro, remains to be overcome.
Methods
In the present study, chicken oviduct epithelial cells (cOECs) were isolated from oviduct tissue and shortly cultured with keratinocyte complete medium supplemented with chicken serum. The isolated cells were characterized with immunofluorescence, western blot, and flow cytometry using oviduct-specific marker. Chicken mutated ovalbumin promoter (Mut-4.4-kb-pOV) was validated in these cells using luciferase reporter analysis.
Results
The isolated cOECs revealed that the oviduct-specific marker, ovalbumin protein, was clearly detected by immunofluorescence, western blot, and flow cytometry analysis revealed that approximately 79.40% of the cells contained this protein. Also, luciferase reporter analysis showed that the constructed Mut-4.4-kb-pOV exhibited 7.1-fold (p<0.001) higher activity in the cOECs.
Conclusion
Collectively, these results demonstrate the efficient isolation and characterization of cOECs and validate the activity of the constructed ovalbumin promoter in the cultured cOECs. The in vitro validation of the recombinant promoter activity in cOECs can facilitate the production of efficient transgenic chickens for potential use as bioreactors.

Keywords: Chicken; Oviduct Epithelial Cells; Ovalbumin Promoter; Luciferase