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Optimization of genetic constructs for high-level expression of the darbepoetin gene in mammalian cells

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Abstract

Genetic constructs were designed in order to optimize darbepoetin production in CHO cells. They are characterized by a higher level of structural optimization of the darbepoetin gene, a higher gene dose, and the selection of promoter elements that have never been used before for this purpose. A transient transfection of CHO cells by the obtained constructs was performed. It was shown that each of the variable factors in the constructs influenced darbepoetin gene expression. A construct containing a doubled dose of the darbepoetin synthetic gene with optimized codon composition under the control of the CMV-EF1α chimerical promoter was proved to be the most efficient.

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Abbreviations

EIA:

enzyme immunoassay

PCR:

polymerase chain reaction

CHO:

Chinese hamster ovary cells

EPO:

erythropoietin

DEPO:

darbepoetin

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Authors and Affiliations

  1. FARMAPARK Company, Moscow, 117246, Russia

    R. R. Shukurov, K. Yu. Kazachenko, A. A. Nurbakov & Yu. A. Seryogin

  2. State Research Institute of Genetics and Selection of Industrial Microorganisms, Moscow, 117545, Russia

    D. G. Kozlov, E. N. Sautkina & R. A. Khamitov

Authors
  1. R. R. Shukurov
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  2. K. Yu. Kazachenko
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  3. D. G. Kozlov
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  4. A. A. Nurbakov
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  5. E. N. Sautkina
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  6. R. A. Khamitov
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  7. Yu. A. Seryogin
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Correspondence to R. R. Shukurov.

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Original Russian Text © R.R. Shukurov, K.Yu. Kazachenko, D.G. Kozlov, A.A. Nurbakov, E.N. Sautkina, R.A. Khamitov, Yu.A. Seryogin, 2013, published in Biotekhnologiya, 2013, No. 2, pp. 34–45.

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Shukurov, R.R., Kazachenko, K.Y., Kozlov, D.G. et al. Optimization of genetic constructs for high-level expression of the darbepoetin gene in mammalian cells. Appl Biochem Microbiol 50, 802–811 (2014). https://doi.org/10.1134/S0003683814090051

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  • DOI: https://doi.org/10.1134/S0003683814090051

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