ABSTRACTIn this study, we isolated and characterized HSP70 cDNA from pufferfish (Takifugu rubripes). The 3053 bp full-length TrHSP70 sequence consisted of a 167 bp 5′-UTR (untranslated region), a 2535 bp open reading frame, and a 351 bp 3′-UTR. BLAST analysis revealed that the TrHSP70 shared high similarity with HSP70 sequences in other species. In our study, we set 3 experimental groups as H1 group (20 °C), H2 group (24 °C), and H3 group (28 °C) for checking the expression level of TrHSP70 in T. rubripes. Tissue-specific gene expression results showed that TrHSP70 had higher expression in the intestines than other tissues of the T. rubripes by RT-qPCR. In the experimental group, we found that the expression of TrHSP70 was upregulated in different tissues in the H3 group. The results show that TrHSP70 is a constitutively expressed gene, which plays an important role in maintaining normal physiological function and coping with stress.