Production of pB602L in Escherichia coli
The B602L gene of ASFV (Pig/HLJ/2018) was synthesized and cloned into pGEX-6p-1 vector by Genscript Biotech Corporation (Nanjing, Jiangsu, China). The recombinant plasmid pGEX-6p-1was transformed into E. coli BL21(DE3) (Qiagen, Hilden, Germany) for pB602L expression. The cells were cultured at 37℃ in LB medium containing 100 μg/ml ampicillin and transferred to 18℃ when the optical density at 600 nm (OD600) reached 0.5~0.6, then the protein expression was induced by 0.3 mM IPTG for 16 h. The cells were lysed using an ultrasonic crusher and then centrifuged for 40 min at 4℃ to remove the celluar debris. Glutathione Sepharose 4B (GE Healthcare, Uppsala, Sweden) was used to purify the GST-tagged pB602L protein, which was then treated with PreScission Protease to remove the GST taggs. The purified pB602L protein was abtained by passing through anion-exchange resin with Resource Q (GE Healthcare, Uppsala, Sweden), and identified by SDS-PAGE and Western-blot.