Document Type : Original Article (s)
Authors
1
Department of Anatomy and Molecular Biology, School of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran
2
Assistant Professor, Department of Genetics and Molecular Biology, School of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran
3
Assistant Professor, Department of Anatomical Sciences, School of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran
4
Department of Biology, School of Sciences, University of Isfahan, Isfahan, Iran
5
Associate Professor, Department of Epidemiology, Isfahan University of Medical Sciences, Isfahan, Iran
Abstract
Background: Improvement of mRNA stability and therefore, production of interferon beta (IFNb) in Chinese hamster ovary (CHO) cell, as a recombinant protein expression system, is very important. The aim of this study was to investigate this effect by creating a codon bias change in instability sequence located in C-terminal of interferon beta protein (Coding region instability determinant or CRID). Methods: Mutations were designed in 4 points, according to codon bias table of Cricetulus griseus, using polymerase chain reaction/splicing by overlapping extension (SOEing PCR) method inside the CRID of interferon beta gene. The concentration of interferon beta in the medium of Chinese hamster ovary cell was determined using ELISA test kit. Findings: The mutations were verified by gel-electrophoresis and sequencing the resultant DNA. The mutations in resulted in a significant (2.9 times) increase in interferon-b concentration (P < 0.05). Conclusion: The mutations in CRID could significantly increase the stability of mRNA and the resultant Interferon b concentration in the medium. These mutations were novel and their effectiveness was proved. Keywords: Interferon beta, Polymerase chain reaction/splicing by overlapping extension (SOEing PCR), C-terminal of interferon beta protein (CRID), Codon bias, Chinese hamster ovary
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