Cloning and expression of the NS3 gene. (A) NS3 gene was amplified from JEV-infected cells by RT-PCR and checked by agarose gel electrophoresis. Lane M, DNA ladder. (B) The recombinant plasmid expressing the C-terminal region of the NS3 protein (NS3c) was double digested with EcoR I and Sal I and separated by agarose gel electrophoresis. Lane M, DNA ladder; Lane Vec, pET-28a empty vector; Lane NS3c, recombinant plasmid expressing NS3c protein. (C) Analysis of the expression and purification of His-tagged NS3c (His-NS3c). Bacterial lysates and purified His-NS3c were separated on a 10% SDS-PAGE gel and stained with Coomassie blue R250. Lane 1, uninduced bacterial cells; lane 2, bacterial cells induced with isopropyl-β-D-thiogalactopyranoside (IPTG); Lane 3, His-NS3c eluted from the His Bind Resin column.