Isopropyl beta d thiogalactopyranoside iptg

chIl-9Rα gene with only extracellular domain was cloned into pET-28a plasmid using a pair of primers (Forward:5′-CGGGATCC(BamHI)AGAGATTTCCCTGGCAGT-3′ and Reverse:5′-CCGCTCGAG(XhoI)CGTTCTGGTGTCAAAGAG-3′) and transformed into E. coli ROSETTA (DE3) strain for recombinant expression by inducing with 0.1 mM isopropyl-beta-D-thiogalactopyranoside (IPTG) (Solarbio, Beijing, China). After induction for 4 h, the lysates of bacteria were analyzed by 12% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and stained with Coomassie Blue (Beyotime, Shanghai, China). In order to produce adequate rchIL-9Rα, 300 mL bacteria culture was induced at the above conditions. The bacteria pellets were sonicated, and the lysed precipitate were dissolved with 8 M urea. The rchIL-9Rα with his tag was purified with the Ni Sepharose Column (General Electric, Boston, MA) according to the manufacturer's protocol. The purified rchIL-9Rα was refolded by dialysis with gradient urea buffer (6 M, 4 M, 2 M, 1 M, 0 M) at 4°C, and finally quantified by BCA Protein Assay Kit (Beyotime, Shanghai, China).