Abstract: To clarify the functions of 3-dehydroquinate dehydratase and shikimate dehydrogenase(DHQ-SDH) genes in Nicotiana tabacum, AtDHQ-SDHs sequences were used to blast in China Tobacco Genome Database and NtDHQ-SDHs were obtained. Gene structure, protein characteristics, cis-regulatory elements and phylogenetic relationships were analyzed by bioinformatics software. Relative expression levels of NtDHQ-SDHs in different tissues of cv. Honghuadajinyuan were detected by qPCR. The results showed that eight NtDHQ-SDHs were identified. The coding region sequences of eight NtDHQ-SDHs were 1 248-1 608 bp in lengths and encoded 415-535 amino acids. The molecular weights of eight NtDHQ-SDHs were 45.13-58.22 kD and the predicted NtDHQ-SDHs were stable proteins. The number of exons and introns in NtDHQ-SDHs were relatively conserved, including 8-10 exons and 7-9 introns. Motif composition and phylogenetic analysis indicated that eight NtDHQ-SDHs were divided into two subfamilies, deriving from N. sylvestris and N. tomentosiformis, respectively. NtDHQ-SDHs were expressed in roots, stems, leaves and terminal buds.