Biotechnology and Bioprocess Engineering 2024; 29(6): 1025-1033  
ROSA26 BAC-based system enables continuous high-yield protein production
Myeong Uk Kuk 1 · Moon Kyoung So 1 · Ji Ho Park 1 · Jee Hee Yoon 1 · Yoo Jin Lee 1 · Duyeol Kim 1 · Byeonghyeon So 1 · Yun Haeng Lee 1 · Minseon Kim 1 · Youngjoo Byun 3 · Hyung Wook Kwon 1,2 · Joon Tae Park 1,2
1 Division of Life Sciences, College of Life Sciences and Bioengineering , Incheon National University , Incheon 22012 , Korea
2 Convergence Research Center for Insect Vectors , Incheon National University , Incheon 22012 , Korea
3 College of Pharmacy , Korea University , Sejong 30019 , Korea
Correspondence to: ✉ Hyung Wook Kwon
hwkwon@inu.ac.kr

✉ Joon Tae Park
joontae.park@inu.ac.kr
Received: May 31, 2024; Revised: September 25, 2024; Accepted: October 8, 2024; Published online: November 15, 2024.
© The Korean Society for Biotechnology and Bioengineering. All rights reserved.

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract
The mouse ROSA26 locus serves as a port to maintain stable expression of the transgene. Therefore, a bacterial artificial chromosome (BAC) containing the ROSA26 locus was used as a platform to produce proteins by incorporating transgenes with foreign promoters. However, the endogenous promoter of the ROSA26 locus, which allows stable high-level transgene expression, has not been applied to the ROSA26 BAC-based platform. In this study, we generated recombinant ROSA26 BAC by targeting “exon 1” or “intron 1” of the ROSA26 locus. Recombinant ROSA26 BAC (exon 1) had lower protein productivity compared to the control. However, recombinant ROSA26 BAC (intron 1) produced protein at a higher efficiency than the control group. The protein productivity induction effect by recombinant ROSA26 BAC (intron 1) was maintained up to a single clone, enabling stable protein production over a long period of time. Taken together, we established a ROSA26 BAC-based protein production system capable of producing protein at high yield over a long period of time.
Keywords: ROSA26 BAC · Endogenous promoter · Intron 1

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