A Heat shock factor (Hsf) from Rhododendron calophytum was cloned using RT-PCR and rapid amplification of cDNA ends (RACE) techniques and it was named RcHsfB3 (accession no. KU145694). The full length was 1130bp, including an open reading frame of 774bp and encoding a protein of 257 amino acids. Its molecular weight and theoretical isoelectric point were 28.8 kDa and 5.38 respectively. Phylogenetic analysis showed that this gene was classified as the same branch with HsfB3 of other plants, indicating that it belongs to the HsfB3 gene family. Multiple sequence alignment analysis indicated that the homology reaches 67.29% with VvHsfB3, which is highest. The result of subcellular localization in protoplasts exhibited that RcHsfB3 is located in the cell nucleus. The expression of RcHsfB3 were examined using real-time polymerase chain reaction. Under heat、cold and salt stress respectively, RcHsfB3 expression level showed an alternate trend of ‘rising and falling’, remarkably up-regulated.Preliminary studies indicate that RcHsfB3 will be induced by high temperature, low temperature and high salt stress.