Establishment and expression verification of serum-free suspension domestication of CHO-K1 monoclonal cell line
10.13200/j.cnki.cjb.004276
- VernacularTitle:无血清悬浮驯化CHO-K1单克隆细胞株的建立及其表达验证
- Author:
XIE Tao
- Publication Type:Journal Article
- From:
Chinese Journal of Biologicals
2024;37(8):945-951
- CountryChina
- Language:Chinese
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Abstract:
ObjectiveTo select a host cell line with traceable monoclonal origin and good growth status by serum-free suspension domestication culture of CHO-K1 cells,and verify its expression.MethodsAdherent CHO-K1 cells were domesticated into suspension cells suitable for serum-free medium by gradually decreasing serum concentration. Monoclonal cells were selected by cell seeding and imaging system Verified In-Situ Plate Seeding(VIPS system for short),and the parameters such as doubling time,agglomeration rate and cell diameter of monoclonal cells were comprehensively evaluated in continuous culture to determine the candidate clones. The candidate clones were transfected with plasmids contained green and red fluorescent proteins respectively,and one clone was identified as the final host cell of the engineering cell line according to the expression of fluorescent proteins.ResultsThe obtained CHO-K1 cell lines were adapted to serum-free suspension culture,and the doubling time was 20-24 h. The monoclonal cell line identified by monoclonal screening and yield evaluation had clear monoclonal origin and could be traced back. At an inoculation density of 0. 5 × 10~6cells/mL,the maximum growth density reached 8. 27 × 10~6 cells/mL through continuous cultivation of 7 d,the viability was not lower than 80% with the mean doubling time of 20. 31 h,and the exogenous gene was effectively expressed.ConclusionThrough serum-free domestication culture,CHO-K1 cell line with traceable monoclonal origin,good growth status and high protein expression was successfully obtained.
