| 摘要 |
Disclosed is a method for highly expressing a recombinant protein in recombinant bacteria, wherein the recombinant protein comprises a hydrophilic and a hydrophobic end, wherein the hydrophilic end is a colicin polypeptide and wherein the hydrophobic end is a polypeptide target moiety which is capable of binding a target, the method comprising: (1) transfecting a recombinant plasmid expressing the recombinant protein into E.coli bacteria with pET system to obtain positive monoclonal colonies and produce recombinant E.coli bacteria, (2) producing seed bacteria solution from the positive monoclonal colonies, and inducing protein expression and enlargement culturing of the seed bacteria solution in enlargement culturing medium; wherein the supernatant of the enlargement cultured solution contains the expressed recombinant protein, (3) extracting and purifying the recombinant protein from the supernatant, wherein the recombinant E.coli bacteria with pET system is E.coli B834 (DE3), and wherein the enlargement culturing medium has water as solvent and comprises the following components: NaCl 6.0~6.7 g/L, peptone 25.0 g/L, yeast powder 7.5 g/L, glucose 0.6~2.0 g/L, Na2HPO4·7H2O 6.8~18.3 g/L, KH2PO4 3.0~4.3 g/L, NH4Cl 1.0~1.4 g/L, MgSO4 0.2~0.4 g/L, CaCl2 0.01 g/L, and methionine 0~40 mg/L. Also disclosed is a medium for recombinant E. Coli bacteria with pET system. wherein the medium has water as solvent and comprises the following components: NaCl 6.0~6.7 g/L, peptone 25.0 g/L, yeast powder 7.5 g/L, glucose 0.6~2.0 g/L, Na2HPO4·7H2O 6.8~18.3 g/L, KH2PO4 3.0~4.3 g/L, NH4Cl 1.0~1.4 g/L, MgSO4 0.2~0.4 g/L, CaCl2 0.01 g/L, and methionine 0~40 mg/L. |
