Objective:To express methyl-CpG-binding domain(MBD) recombinant proteins in E. coli. Methods:DNA encoding MBD of human methyl-CpG-binding protein 2(Mecp2) was optimized for E. coli preferred codons. The codon-optimized MBD coding sequence was synthesized,and then was subcloned into pGS21a to construct recombinant expression plasmid. Recombinant protein expression was induced in E. coli Rosseta-gami(DE3). Recombinant proteins were detected by SDS-PAGE and Western blot,and then purified using Nickel-affinity chromatography columns. The interaction between recombinant MBD proteins and methylated DNA was analyzed by surface plasmon resonance(SPR). Results:Enzyme digestion and DNA sequencing demonstrated the prokaryotic expression vector carrying codon-optimized MBD gene was successfully constructed. SDS-PAGE and Western blot results indicated that recombinant MBD proteins were expressed in E. coli. Finally,recombinant MBD proteins with the relative molecular weight of 38 000 were purified by affinity chromatography. SPR analysis demonstrated that the recombinant MBD proteins could bind methylated DNA specially. Conclusion:The prokaryotic expression vector carrying codon-optimized MBD gene was successfully constructed,and recombinant MBD proteins were expressed in E. coli scucessfully.