In vitro replication system mediated by NIb of Potato virus Y

Abstract

Potato virus Y (PVY) is a serious global virus. At present, PVY are mainly focused on strain analysis and protein interaction with host, but independent studies on PVY replication in vitro are still weak, the main problem is that there is no independent in vitro replication system for PVY. This study was focused on the creation of in vitro replication system mediated by NIb of PVY. The optimal expression concentration of MBP-NIb is 0.2 mM by the IPTG gradient experiment. Based on the solubility analysis, it is suggested that 37℃ is feasible and the soluble ratio of MBP-NIb is about 10%, which is enough for following affinity chromatography against MBP-tag. The purified MBP-NIb can specifically recognize the 3' terminal region of PVY plus or minus strand and perform in vitro replication. This is the first time that an in vitro replication system for PVY has been established. This in vitro replication system will facilitate the study of mechanism on PVY replication.