Comparison of Vero and MDCK cell lines transfected with human siat7e gene for conversion to suspension culture

Abstract

Introduction: Inactivated influenza vaccines are traditionally produced in chicken embryonated eggs but its limitations in producing the required doses in pandemic outbreaks quickly enough has made searching for alternative modes of production necessary. The use of cell culture-based vaccine production is one way of overcoming the limitations of the egg-based method and securing a more rapid response. Although Vero cells are suitable for production of influenza vaccine, but their anchorage-dependency limits their production capability. In this study adherent Vero and MDCK cells were transfected with human siat7e gene in order to convert anchorage-dependent cells to those capable of growing in suspension. Methods: Human siat7e gene was amplified with primers containing restriction sites for Xho I and Hind III and the product was cloned into pEGFP-N1 vector upstream of GFP sequence. The cells were transfected with the construct containing the siat7e gene and a medium containing G418 was used to select stably transfected cells which were then evaluated using inverted immunofluorescence microscopy. Results: Anchorage-dependent cells exhibited changes in cell-cell adhesion and cell spreading behavior following transfection. Vero cells showed a higher longevity compared to MDCK cells as viability of the latter cells declined after 50 h. Conclusion: The data showed that adherent Vero cells can successfully be converted to anchorage- independent cells capable of growing in suspension through transfection with human siat7e gene.

Keywords

• human siat7e
• influenza virus
• transfection
• vero cell
• suspension culture.