Fig. 4.

Fig. 4. Cloning and heterologous expression of T26gal. (A) Schematic presentation of the pET-T26gal plasmid; (B) Cloning of the T26gal gene (M: DM0004 DNA Marker; 1: T26gal gene; CK: blank control); (C) Restriction enzyme digestion verification of pET-T26gal (M: DM0005 DNA Marker; 1: Digestion by XhoI; 2: Digestion by NdeI; 3: Double digestion by XhoI/ NdeI; (D) IPTG-induced expression of T26GAL in E. coli BL21 (M: Standard molecular weight of protein 1: Total protein of BL21/pET26b(+) before IPTG induction, 2: Total protein of BL21/pET26b(+) after IPTG induction, 3: Total protein of BL21/ pET-T26gal before IPTG induction, 4: Total protein of BL21/pET-T26gal after IPTG induction, the arrow indicates the target protein. (E) Ni-affinity column chromatography purification of the recombinant α-galactosidase T26GAL (M: Standard molecular weight of protein, 1 and 2: The extraction of IPTG induced BL21/pET26b(+) cell pellets, 3: the sample after passing Ni-affinity column, 4 and 5: the washout of the Ni-affinity column with 100 mM imidazole, 6 and 7: the washout of the Ni-affinity column with 200 mM imidazole, 8 and 9: the washout of the Ni-affinity column with 300 mM imidazole).

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