Figure 3 | Scientific Reports

Figure 3

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

Figure 3

Transfer of the MAR HAC vector into CHO K1 cells promotes protein production. (A) Schematic diagram of transfer of the HAC vector in CHO DG44 cells to CHO K1 cells via MMCT. (B) Microcell formation was induced by colcemid treatment for 72 hr in CHO DG44 cells. Scale bars: 50 μm. (C) Two-color FISH analysis of the HAC vector in G418-resistant CHO K1 clones was performed with digoxigenin-labeled hCot-1 DNA (red) and biotin-labeled EGFP (green). The arrows indicate large metacentric chromosomes that identify the CHO strain. The left panel shows CHO K1R EGFP MAR-HAC cl.1. The right panel shows CHO K1R EGFP MAR-HAC cl.2. The arrowhead indicates the HAC, and the inset shows enlarged images of the HAC (EGFP probe-specific green signals are indicated by the small arrowhead). (D) Expression levels of EGFP were analyzed with western blotting. CHO DG44 MAR-HAC cl.1 (Donor cells) was used as a control. Expression levels of EGFP were normalized to levels of tubulin. The expression level of protein in DG44 EGFP MAR-HAC cl.1 was arbitrarily assigned as 1. Cropped blots were used in this figure. Original full-length blots are presented in Supplementary Fig. S3. Data shown are representative of at least three independent experiments.

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