Identification of functional cis-acting RNA elements in the hepatitis E virus genome required for viral replication
Fig 5
Full-length HEV genome with synonymous mutations in the cis-acting RNA elements is significantly impaired in its ability to produce infectious virus.
(A) Schematic diagrams of the protocol to determine the production of infectious virus. For the cis-acting element mutants, synonymous mutations were introduced in the ORF1 coding region (G113C or G113T) or ORF2 coding region (G7335A). (B-C) Transfection of in vitro-transcribed WT, synonymous mutants or GAD (Pol-) RNA of full-length Kernow C1/p6 (GT 3) into S10-3 cells. Cell lysate supernatant was collected 7 days after transfection, and virus was titrated by infecting HepG2C3A cells. Cells were stained with anti-HEV ORF2 mAbs at 5 days post of infection. (B) to quantify infectious viral particles by foci-forming assay (C). Values are means plus SD (n = 3). IF, immunofluorescence; FFU, foci-forming units; LOD, limit of detection.