Chymopapain gene（CHY） was cloned from papaya by RT-PCR, and constructed into pEASY-E1 togenerated recombinant plasmid pEASY-E1-CHY. The plasmid was transformed into E.coli. The expression was in-duced by IPTG and the induction time was optimized. The condition of suitable expression was 6h and 0.05 mmlo/LIPTG. The analysis of SDS-PAGE and Western-Blot indicated that the molecular weight of protein was 45 kDa. Thechymopapain was expressed successfully in BL21（DE3）and the enzyme activity was identified by curd experiment.The result will be helpful to the chymopapain engineering.