The prokaryotic expression vectors pET-EuCHIT1 and pMCSG-EuCHIT1 of Eucommia ulmoides chitinase gene ( EuCHIT1) were constructed with strong T7 promoter. The expression products of pET-EuCHIT1 and pMCSG-Eu-CHIT1 contained six histidine (6His) tags and six maltose-binding proteins (his6-tag-maltose-binding protein, MBP), respectively. The recombinant vectors were transformed into E.coli cell strain BL21(DE3) for expression. The transformed cell were cultured under the conditions of 37℃ and 150 r until OD value of the bacteria was 0.4~0.8, then induced for 12 h by 1 mmol/L IPTG under the conditions of 16℃ and 150 r. The expression proteins were analysed by SDS-PAGE. The expression of pET-EuCHIT1/BL21(DE3) contained 36.03 kD fusion protein, and pMCSG-EuCHIT1/BL21(DE3) DE3) successfully expressed the soluble 77. 21 kD fusion protein. It is concluded that the chtinase gene ( EuCHIT1) could be expressed in pMCSG-EuCHIT1/BL21( DE3) to obtain soluble fusion protein, thus laying a foundation for prepa-ration of EuCHIT1 polyclonal antibody and for study of its function.
EuCHIT1Prokaryotic expressionSoluble fusion protein
NETL
NSTL
维普
万方数据
