1. Academic Validation
  2. An efficient protein production system via gene amplification on a human artificial chromosome and the chromosome transfer to CHO cells

An efficient protein production system via gene amplification on a human artificial chromosome and the chromosome transfer to CHO cells

  • Sci Rep. 2019 Nov 18;9(1):16954. doi: 10.1038/s41598-019-53116-2.
Takahito Ohira 1 2 Koichi Miyauchi 1 Narumi Uno 1 2 3 Noriaki Shimizu 4 Yasuhiro Kazuki 1 2 Mitsuo Oshimura 2 Hiroyuki Kugoh 5 6
Affiliations

Affiliations

  • 1 Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, Yonago, Tottori, 683-8503, Japan.
  • 2 Chromosome Engineering Research Center, Tottori University, Yonago, Tottori, 683-8503, Japan.
  • 3 Department of Applied Life Sciences, Tokyo University of Pharmacy and Life Sciences, Horinouchi, Hachioji, Tokyo, 192-0392, Japan.
  • 4 Graduate School of Biosphere Science, Hiroshima University, Higashi-hiroshima, Hiroshima, 739-8521, Japan.
  • 5 Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, Yonago, Tottori, 683-8503, Japan. kugoh@med.tottori-u.ac.jp.
  • 6 Chromosome Engineering Research Center, Tottori University, Yonago, Tottori, 683-8503, Japan. kugoh@med.tottori-u.ac.jp.
Abstract

Gene amplification methods play a crucial role in establishment of cells that produce high levels of recombinant protein. However, the stability of such cell lines and the level of recombinant protein produced continue to be suboptimal. Here, we used a combination of a human artificial chromosome (HAC) vector and initiation region (IR)/matrix attachment region (MAR) gene amplification method to establish stable cells that produce high levels of recombinant protein. Amplification of Enhanced green Fluorescent protein (EGFP) was induced on a HAC carrying EGFP gene and IR/MAR sequences (EGFP MAR-HAC) in CHO DG44 cells. The expression level of EGFP increased approximately 6-fold compared to the original HAC without IR/MAR sequences. Additionally, anti-vascular endothelial growth factor (VEGF) antibody on a HAC (VEGF MAR-HAC) was also amplified by utilization of this IR/MAR-HAC system, and anti-VEGF antibody levels were approximately 2-fold higher compared with levels in control cells without IR/MAR. Furthermore, the expression of anti-VEGF antibody with VEGF MAR-HAC in CHO-K1 cells increased 2.3-fold compared with that of CHO DG44 cells. Taken together, the IR/MAR-HAC system facilitated amplification of a gene of interest on the HAC vector, and could be used to establish a novel cell line that stably produced protein from mammalian cells.

Figures
Products
  • Cat. No.
    Product Name
    Description
    Target
    Research Area
  • HY-P9906
    ≥99.0%, VEGF Blocking Antibody