Abstract: In order to investigate the molecular biological characteristics of the poultry-derived double-stranded RNA-dependent protein kinase（PKR） gene and to understand its expression distribution in chicken tissues/organs, in this experiment, firstly, the physical and chemical properties of the coding protein were analyzed and the secondary structure and tertiary structure were predicted using the bioinformatics online tool. The poultry-derived PKR gene was then inserted into the pEF1α-Myc eukaryotic expression vector; the recombinant eukaryotic expression vector pEF1α-Myc-PKR was then transfected into DF1 cells. The expressed protein were verified by indirect immunofluorescence and Western-blot identification, and subcellular localization was performed by laser confocal scanning microscopy. Finally, real-time PCR was used to detect the distribution of the gene in different tissues/organs of SPF chicken. The results showed that the molecular formula of poultry-derived PKR protein was C₂₇₄₄H₄₃₃₂N₇₇₂O₈₄₂S₂₃, and a total of 550 amino acids were encoded. The theoretical molecular mass was 62 346.63 u, the isoelectric point（pI） was 8.63, and the instability coefficient was 45.54, which was an unstable protein. Among the secondary structure, the proportion of α-helix was 33.27%, the proportion of extended chain was 13.64%, the proportion of β-corner was 4.73%, and the proportion of irregular curl was 48.36%. The similarity between the tertiary structure prediction model and human PKR protein was 47.29%. The recombinant eukaryotic expression vector pEF1α-Myc-PKR was successfully constructed by double digestion, and the recombinant eukaryotic expression vector pEF1α-Myc-PKR could express PKR protein in DF1 cells. The molecular mass of the protein was about 62 ku, which had good reactogenicity and was mainly localized in the cytoplasm. Poultry-derived PKR gene was expressed in all 17 tissues/organs of SPF chicken, but the expression was different in different tissues/organs. The highest expression was in the blood, followed by the pancreas, intestines, lung, spleen, thymus, bursa, liver, glandular stomach, muscle stomach, skin, trachea, brain and kidney; extremely low expression was in joints and muscles. The results speculated that this gene might be involved in blood immunity reaction, and played an important role in the innate immune response.