Cloning and Expression of TspGST Protein

The TspGST gene (GenBank accession no. XM_003373603) was amplified by PCR using specific primers with BamHI and Hind III restriction enzyme sites (underlined) (forward: 5′-TAT AGG ATC CAT GAC CAA CAC GTC GAA GAA AGG-3′; reverse, 5′-GCC CAA GCT TTC ATT GAC TTT CAA TAG TCA CTG G-3′). The purified PCR product was cloned into the pMD19-T vector (Takara, Dalian, China), subsequently sub-cloned into the pQE-80 L (Novagen, La Jolla, CA, USA). The recombinant plasmid carrying the TspGST gene was transformed into Escherichia coli BL21 (DE3) (Novagen), and expressed under IPTG induction. The rTspGST was purified using Ni-NTA-Sefinose resin (Sangon Biotech, Shanghai, China). The concentration of the purified rTspGST was assayed as described previously [22 (link)], and identified by SDS-PAGE analysis [23 (link)]. The gel was stained with 0.25% Coomassie brilliant blue R-250 (Sigma-Aldrich), and subsequently decolorized.