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Comparative Assessment on the Expression Level of Recombinant Human Follicle-Stimulating Hormone (FSH) in Serum-Containing Versus Protein-Free Culture Media

Author:
Jazayeri, Seyedeh Hoda, Amiri-Yekta, Amir, Gourabi, Hamid, Abd Emami, Baharak, Halfinezhad, Zahra, Abolghasemi, Somayeh, Fatemi, Nayeralsadat, Daneshipour, Abbas, Ghahremani, Mohammad Hossein, Sanati, Mohammad Hossein, Khorramizadeh, Mohammad Reza
Source:
Molecular biotechnology 2017 v.59 no.11-12 pp. 490-498
ISSN:
1073-6085
Subject:
Chinese hamsters, Western blotting, biopharmaceuticals, biotechnology, cell lines, culture media, dihydrofolate reductase, enzyme-linked immunosorbent assay, follicle-stimulating hormone, gene amplification, gene expression, humans, messenger RNA, protein synthesis, quantitative polymerase chain reaction
Abstract:
Production of recombinant pharmaceutical proteins has made a great contribution to modern biotechnology. At present, quick advances in protein expression lead to the enhancement of product quantity and quality as well as reduction in timescale processing. In the current study, we assessed the expression level of recombinant human follicle-stimulating hormone (rhFSH) in adherent and suspension Chinese hamster ovary (CHO) cell lines by cultivation in serum-containing and chemically defined, protein-free media. The expression cassette entailing FSH subunits was transfected to CHO/dhfr- and CHO DG44 cell lines, and gene amplification was achieved using dihydrofolate reductase (DHFR)/methotrexate (MTX) system. Afterward, the expression level of rhFSH was studied using real-time PCR, Western blotting and ELISA. Our achievements revealed that stepwise increase in MTX [up to 2000 nano-molar (nM)] leads to boost the expression level of rhFSH mRNA in both cell lines, although DG44 have better results, as mRNA expression level reached 124.8- and 168.3-fold in alpha and beta subunits, respectively. DG44 cells have also the best protein production in 2000 nM MTX, which reached 1.7-fold in comparison with that of the mock group. According to the above results and many advantages of protein-free media, DG44 is preferable cell line for future steps.
Agid:
5844441