Ni nta 6ff sefinose resin kit

Each of the supernatant solutions containing the expressed proteins were collected by means of centrifugation at 4000× g for 5 min and were directly applied to the dialysis membranes (10 kDa). With respect to protein concentration, we covered the dialysis bag with PEG 20,000 solid dispersions. When the volume was concentrated to 10 mL, the solutions were purified using the Ni-NTA 6FF Sefinose™ Resin Kit (Sangon Biotech, Shanghai, China). Finally, the purified proteins were concentrated with the Amicon^® Pro System (Merck, Kenilworth, NJ, USA). The purity of these proteins was analyzed by the SDS-PAGE system.

After sequencing to confirm that it is correct, the expression of recombinant RIPK3 truncated protein was induced at 37 °C and 20 °C, respectively, adding 1 mM isopropyl 1-thio-beta-d-galactopyranoside (IPTG) into liquid LB/Amp medium. Expression levels of the induced RIPK3 were identified with 12% SDS-PAGE and purified with Ni-NTA 6FF Sefinose Resin Kit (Sangon Biotech, Shanghai, China). Subsequently, Western blot analysis with anti-His tag further verified that the protein purified is the target protein containing His-tag. Finally, the target protein eluted by the optimal concentration of imidazole was subjected to dialysis and renature.