pMH3-rFVII

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	The constructed plasmids were transfected to CHO K1 cells by electroporation using 400V for 40 μs. The electroporation reaction mixture contained 3×106 cells, 20 μg of plasmid, and 5 μg of salmon sperm DNA. The transfected cells were cultured in DMEM/F 12 with 10% fetal bovine serum (FBS) and selected with 2.0 mg/ml G418 at 37°C, 5% CO2 for 48 h.	The surviving clones were picked and cultured in a 96-well microplate with DMEM/F 12 supplemented with 10% FBS. After culturing the sub-clones for 6 d, the cells were cultured in M1 SFM for 2 d. The supernatant from each well was analyzed by dot blot and high expression clones were subsequently cultured in a 6-well microplate. High expression clones in 6-well microplates were screened by protein gel blot (WB) and the subsequent high expression sub-clones were selected for further suspension culture.

Protocol tips
The constructed plasmids were transfected to CHO K1 cells by electroporation using 400V for 40 μs. The electroporation reaction mixture contained 3×106 cells, 20 μg of plasmid, and 5 μg of salmon sperm DNA. The transfected cells were cultured in DMEM/F 12 with 10% fetal bovine serum (FBS) and selected with 2.0 mg/ml G418 at 37°C, 5% CO2 for 48 h.
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The surviving clones were picked and cultured in a 96-well microplate with DMEM/F 12 supplemented with 10% FBS. After culturing the sub-clones for 6 d, the cells were cultured in M1 SFM for 2 d. The supernatant from each well was analyzed by dot blot and high expression clones were subsequently cultured in a 6-well microplate. High expression clones in 6-well microplates were screened by protein gel blot (WB) and the subsequent high expression sub-clones were selected for further suspension culture.

Publication protocol

The constructed plasmids were transfected to CHO K1 cells by electroporation using 400V for 40 μs. The electroporation reaction mixture contained 3×106 cells, 20 μg of plasmid, and 5 μg of salmon sperm DNA. The transfected cells were cultured in DMEM/F 12 with 10% fetal bovine serum (FBS) and selected with 2.0 mg/ml G418 at 37°C, 5% CO2 for 48 h. The surviving clones were picked and cultured in a 96-well microplate with DMEM/F 12 supplemented with 10% FBS. After culturing the sub-clones for 6 d, the cells were cultured in M1 SFM for 2 d. The supernatant from each well was analyzed by dot blot and high expression clones were subsequently cultured in a 6-well microplate. High expression clones in 6-well microplates were screened by protein gel blot (WB) and the subsequent high expression sub-clones were selected for further suspension culture.

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