Abstract: The cathepsin L cDNA of Penaeus monodon (PmCatL) was cloned by rapid amplification of cDNA ends (RACE). The PmCatL cDNA included an open reading frame (ORF) of 1 026 bp encoding a polypeptide of 341 amino acids, which contained a typical signal peptide sequence (Met¹-Ala¹⁸), a prodomain (Val¹⁹-Gln¹²³) and a mature domain (Leu¹²⁴-Val³⁴¹). The preprocathepsin contained a potential N-glycosylation site (Asn⁹⁹), three catalytic sites (Cys¹⁴⁸, His²⁸⁷ and Asn³⁰⁸)and sequences E⁴⁵X₃RX₃F/WX₂NX₃IX₃N⁶⁴ and G¹⁸⁷CXGG¹⁹² were discovered in amino acid sequence. Homology analysis of PmCatL reveals that the PmCatL shared 36%~75% identity to other known cathepsin L sequences. The phylogenetic tree shows that the PmCatL clustered with the invertebrate cathepsin L cysteine proteases and was closely related to Macrobrachium rosenbergii cathepsin L. The expression levels of PmCatL in different tissues were analyzed by quantitative real-time PCR. Expression of PmCatL was detected in all tested tissues and the highest expression level was in lymph. The expression levels of PmCatL in different growth periods show that the expression levels of PmCatL with the highest expression level in post-larval, PmCatL might be associated with growth and development of P.monodon. PmCatL had the highest expression at Stage Ⅳ, which suggests that PmCatL might play an important role in ovary development in P.monodon.