Optimization of Genetic Constructs for High-level Expression of Darbepoetin Gene in Mammalian Cells

Автор: R.R. Shukurov, K.Yu. Kazachenko, D.G. Kozlov, A.A. Nurbakov, E.N. Sautkina, R.A. Khamitov, and Yu.A. Seryogin

Страница: 34-45

Optimization of genetic constructs for high-level expression of Darbepoetin Gene in mammalian cells

Biotekhnologiya, 2013, N 2, P. 34-45

UDC 606:57.082.26

Section:  “Producers, Biology, Selection, and Gene Engineering”

R.R. Shukurov ^{1, *},  K.Yu. Kazachenko ¹,  D.G. Kozlov ²,  A.A. Nurbakov ¹,  E.N. Sautkina ², R.A.  Khamitov ²,  and  Yu.A. Seryogin ¹

¹  The Limited Liability Company “PHARMAPARK”,  117246, Moscow Russia

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

e-mail:  anamezon@gmail.com

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

Key words:  CHO cells,  codon optimization,  darbepoetin-alpha,  erythropoetin,  expression vectors,  promoter.

The full English version of the article was published in “Applied Biochemistry and Microbiology”, 2014, Vol. 50, Issue 9, pp. 802-811 as R. R. Shukurov, K. Yu. Kazachenko, D. G. Kozlov, A. A. Nurbakov, E. N. Sautkina, R. A. Khamitov, and Yu. A. Seryogin “Optimization of genetic constructs for high-level expression of the darbepoetin gene in mammalian cells”.

It is contained at the SpringerLink website:  http://link.springer.com/article/10.1134/S0003683814090051

DOI: 10.1134/S0003683814090051

10.04.2015, 1896 просмотров.