High-level expression of human thyroid-stimulating hormone in Chinese hamster ovary cells by co-transfection of dicistronic expression vectors followed by a dual-marker amplification strategy

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作者：

Cibele，N.，Peroni，Carlos，R.，J.，Soares，Elizabeth，Gimbo，Ligia

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摘要：

The utilization of dicistronic mRNA expression vectors, containing the gene of interest upstream of an amplifiable marker gene, has shown success in rapidly, efficiently and reproducibly obtaining stable cell lines that express high levels of the protein of interest. For this reason, human thyroid-stimulating hormone (hTSH), a heterodimeric glycoprotein composed of non-covalently linked α- and β-subunits, was expressed in Chinese hamster ovary (CHO) cells using a system based on dicistronic expression vectors. These contained the genes of interest and the amplifiable gene markers dihydrofolate reductase (DHFR) and adenosine deaminase (ADA), separated by an internal ribosome entry site isolated from the encephalomyocarditis virus. After the cells (CHO-DHFR<sup>−</sup>) had been co-transfected with the expression vectors and submitted to gene amplification in culture medium containing stepwise increments of methotrexate, it was possible to isolate clones that presented a

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关键词：

adenosine deaminase CHO cell dihydrofolate reductase encephalomyocarditis thyrotropin.