New reverse genetics and transfection methods to rescue arboviruses in mosquito cells

Reverse geneticsis a critical tool to decrypt the biological properties of arboviruses. However, whilst reverse geneticsmethods have been usually applied to vertebrate cells, their use in insect cells remains uncommon due to the conjunction of laborious molecular biology techniques and of specific difficulties surrounding the transfection of such cells. To leverage reverse geneticsstudies in both vertebrate and mosquito cells, we designed an improved DNA transfection protocol for insect cells and then demonstrated that the simple and flexible ISA (Infectious Subgenomic Amplicons) reverse- genetics methodcan be efficiently applied to both mammalian and mosquito cells to generate in days recombinant infectious positive-stranded RNA viruses belonging to genera Flavivirus( Japanese encephalitis, Yellow fever, West Nile and Zika viruses) and Alphavirus( Chikungunyavirus). This method represents an effective option to potentially overcome technological issues related to the study of arboviruses.