Abstract: In order to reveal the function and mechanism of nucleoside diphosphate kinase in tobacco, NtNDPK4 was cloned from tobacco cv. K326. The CDS length of NtNDPK4 was 678 bp, and 225 amino acids were encoded. Bioinformatics analysis showed that the molecular weight of NtNDPK4 protein was 24.95 kDa, and the pI was about 8.34. The subcellular localization was predicted in chloroplast. The evolutionary analysis showed that NtNDPK4 belonged to NDPK Ⅱ. The cis-acting element analysis predicted that the promoter of NtNDPK4 contained hormone and light-responsive elements. Tissue expression specificity analysis showed that NtNDPK4 was expressed in all tissues. Under the treatment of different hormones(MeJA, ABA, GR-24, IAA, GA and 6-BA), the expression of NtNDPK4 was significantly up-regulated, and the expression level of NtNDPK4 was up-regulated nearly 50 folds after the GA treatment. After 24 h of light restoration, the expression of NtNDPK4 in tobacco seedlings grown in darkness was also up-regulated obviously. These results suggest that NtNDPK4 might be involved in hormone and light responses.