Overexpression of the recombinant human interferon-beta (rhIFN-β) gene in tobacco chloroplasts

Abstract

Chloroplast genetic engineering is a convenient method for the production of recombinant proteins by increasing the expression level of transgenes. Interferon-beta (IFN-β) is a member of type I interferons that possess some pharmaceutical properties. The present study aimed to investigate the overexpression and production of the recombinant human IFN-β gene (rhIFN-β) in the tobacco chloroplast genome. For this purpose, a codon-optimized rhIFN-β was transferred to the pVSR326 plastid vector containing the aadA gene as a selectable marker. The rhIFN-β gene was then successfully introduced into the tobacco chloroplast genome by using a gene gun. The integration of the rhIFN-β gene into the chloroplast genome and the homoplasmy of the T1 progeny were confirmed by PCR and Southern blot analysis, respectively. RT-PCR and western blot analyses confirmed the transcription and translation of the rhIFN-β gene, respectively. An enzyme-linked immunosorbent assay (ELISA) showed that the rhIFN-β protein in transplastomic plants comprised approximately 2.4% of total soluble protein (TSPs). The bioassay confirmed that the rhIFN-β protein expressed in the tobacco chloroplast had a relatively high biological activity (2.9×104 IU/ml) and protected human amnionic cells against the vesicular stomatitis virus (VSV). On the basis of these findings, it can be concluded that plastid transformation can serve as an operative method for the production of pharmaceutical recombinant proteins.

Keywords

• plastid expression
• chloroplast engineering
• homoplasmy
• pharmaceutical protein
• molecular farming