Patent ID: 8771988
Filing Date: 2014-07-08
Classification: C07K,C12N,C12P

Abstract:
1. A method for the recombinant production of a heterologous immunoglobulin in a CHO cell which is secreted to the cultivation medium comprising: a) providing a CHO cell, wherein said CHO cell is b) providing a nucleic acid comprising whereby a first transfection vector is provided which comprises said provided nucleic acid and an additional fourth nucleic acid sequence conferring resistance to a first eukaryotic selection agent, whereby a second transfection vector is provided which comprises said provided nucleic acid and an additional fourth nucleic acid sequence conferring resistance to a second eukaryotic selection agent, whereby said second eukaryotic selection agent is different to said first eukaryotic selection agent, b1) providing a nucleic acid comprising whereby a third transfection vector is provided which comprises said provided nucleic acid and an additional fourth nucleic acid sequence conferring resistance to a third eukaryotic selection agent, whereby said third eukaryotic selection agent is different to said first eukaryotic selection agent and is also different to said second eukaryotic selection agent, c) transfecting said CHO cell, wherein said transfecting comprises the following: d) cultivating said transfected CHO cell in a medium in the presence of said first and said second and third eukaryotic selection agent, under conditions suitable for the expression of said second, and/or third nucleic acid, and e) recovering said secreted heterologous immunoglobulin from the cultivation medium and thereby producing a heterologous immunoglobulin in a CHO cell which is secreted to the cultivation medium; wherein said resultant CHO cell is stable in the absence of any or all selection agents, as used in the previous steps, for up to generation 60.