Stability study on subculturing of working seed strain for recombinant human interferon α2b
10.13200/j.cnki.cjb.004596
- VernacularTitle:重组人干扰素α2b工作代菌种的传代稳定性
- Author:
Kaining ZHANG
- Publication Type:Journal Article
- Keywords:
Recombinant human interferon(rhIFN) α2b;
Working seed strain;
Passage stability;
Scale-down model
- From:
Chinese Journal of Biologicals
2025;38(11):1306-1312
- CountryChina
- Language:Chinese
-
Abstract:
Objective To verify the passage stability of recombinant human interferon(rhIFN) α2b working seed strain at key generations through the construction of a 15 L scale-down model, in order to ensure the stability and effectiveness of rhIFNα2b during the production process.Methods The rhIFNα2b lyophilized strain was revived and subcultured, and glycerol stocks of the 0, 10 th, 20 th, 30 th, 40 th and 50 th generations of the culture were preserved respectively. The glycerol stocks of each generation preserved in the subculture stability study were selected and the fermentation-expression experiments were conducted in a 15 L reduced-scale model fermentation system under simulated production conditions. By real-time monitoring of the growth curve of the cells and combining with endpoint detection of product expression level,biological activity, plasmid stability, genetic integrity and microbial purity, a comprehensive comparison was made to determine whether the limited subculture times could cover the existing culture process.Results The rhIFNα2b working seed strains from passages 0, 10, 20, 30, 40, and 50 exhibited good stability and consistency after continuous subculturing.All strains showed typical E. coli colony morphology on LB agar plates with no contamination by foreign microorganisms.Antibiotic resistance profiles remained unchanged, Gram staining confirmed Gram-negative bacterial characteristics, and biochemical profiles were consistent with typical E.coli traits. The target gene expression levels consistently exceeded 20%without a declining trend, and the fermentation activity remained above 1. 0 × 108 IU/L. Plasmid restriction analysis produced bands consistent with theoretical expectations, with no observed sequence mutations, and the plasmid loss rates remained below 30%, indicating robust genetic stability.Conclusion The rhIFNα2b working seed strains demonstrated good fermen-tation stability under scaling-down simulated production conditions in the 15L bioreactor. The current passage limitations of the strain effectively meet the requirements of the updated production process, providing crucial data to support process improvement and optimization.
