Abstract: To investigate the potential function of ubiquitin carboxyl terminal hydrolases（UCHs） in the ubiquitination process of laticiferous of Hevea brasiliensis, the full-length sequences of two UCHs family members（HbUCH-L3 and HbUCH-L5） were isolated from Hevea brasiliensis, and both had typical UCHs domains. The open reading frame of HbUCH-L3 and HbUCH-L5 were 993 bp and 558 bp, and encoded 330 and 185 amino acids, respectively. The results of qRT-PCR showed that HbUCH-L3 and HbUCH-L5 were constitutively expressed in all tissues, but was low in latex. In vitro ubiquitination substrates cleavage of recombinant HbUCHs showed that both HbUCH-L3 and HbUCH-L5 had the function of hydrolyzing ubiquitin.HbUCHs significantly reduced the overall ubiquitination level of C-serum proteins; while, the deubiquitinating activity of HbUCH-L3 was higher than that of HbUCH-L5. Therefore, it was speculated that UCHs played a role in maintaining the dynamic balance of laticiferous ubiquitination and thus played a specific biological function, but the exact mechanism was still unclear.