CONSTRUCTION OF AD293-LACI STABLE CELL LINE USING LENTIVIRUS FOR ADENOVIRUS-BASED VACCINEÂ PRODUCTION

• Main Author: Nurfitri Annandari, Afinna
• Format: Theses
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/85322
• Institution: Institut Teknologi Bandung

AD293 cells are a mammalian host cell line used for the production of adenovirusbased vaccines. Production of adenovirus as a vector for vaccines can be suppresed by the expression of vaccine protein coding genes. The Lac system is a transcriptional regulatory system that can be used to inhibit transgene protein expression in host cells. This research aims to construct AD293-lacI stable cell line for the production of adenovirus vector-based vaccines with the lac operon regulatory system. AD293-lacI cells were constructed by infection of lentivirus carrying lacI into AD293 cells. Transduced cells were selected with medium containing 1 ?g/mL puromycin for 12 days. Selected cells were further propagated, meanwhile the negative control was eliminated in the selection medium. The AD293-lacI cells were further used for the AdV_Spike and AdV_LacO_Spike production, using AD293 and AD293-lacI cells (transient) as controls. AdV_Spike and AdV_LacO_Spike titer determination in AD293 were 8.9 × 10 7 and 7.3 × 10 7 IFU/mL. The titer of AdV_Spike and AdV_LacO_Spike produced in AD293-lacI were 1.3 × 10 7 and 1.2 × 10 7 IFU/mL, meanwhile in AD293 cells -lacI (transient) were 1.2 × 10 6 and 2.5 × 10 6 IFU/mL. Detection of Spike protein expression upon AdV_Spike and AdV_LacO_Spike infection showed that the inhibition of Spike protein expression using the lac system was not successful. Although the AD293- lacI was successfully grown in puromycin containing media, the stable expression of lacI protein need to be further analyzed.