Patent ID: 8771983

Claim:
A method of expressing a protein from a repeated sequence formed in CHO cells in which gene amplification is induced, the protein having been under expression repression, the method comprising (1) transfecting into the CHO cells (a) a gene structure comprising (i) a first polynucleotide having an origin of replication and a nuclear matrix attachment region that functions in eukaryotic cells and a selection marker that is a drug resistance gene which confers resistance to an antibiotic, and (ii) a second polynucleotide having a promoter region controllably linked to a polynucleotide encoding a protein to be expressed and an insulator sequence; and (b) a polynucleotide encoding a transcription activation factor of the promoter; and (2) subsequently treating the transfected CHO cells with 5-aza-2′-deoxycytidine at a concentration in the range of 1-2 μM and culturing the transfected CHO cells sequentially in a medium of increasing concentrations of the antibiotic whereby the concentration of the antibiotic increases by 10% to 30% per 24 hours.