Abstract: In order to study the effects of NtPHGPx on salt tolerance of tobacco, NtPHGPx gene overexpression (OX) and CRISPR/Cas9 knockout (Cri) lines were obtained. Two OX and one Cri lines were subjected to 150 mmol/L NaCl treatments for 14 d, and root length, GPx activity, malondialdehyde (MDA) and hydrogen peroxide (H₂O₂) contents were determined. The results showed that the expression of NtPHGPx was induced by NaCl 3 h after treatment. Salt treatments significantly inhibited root growth of wild type tobacco and increased the accumulation of MDA and H₂O₂. Under salt stress, root length of the OX lines was longer than that of wild type and the contents of MDA and H₂O₂ were lower. However, root growth of the Cri line was further inhibited, and the activity of GPx was significantly decreased. The accumulation of MDA and H₂O₂ in shoots was significantly higher than that of wild type. In conclusion, overexpression of NtPHGPx improved, however knockout of NtPHGPx attenuated salt tolerance of tobacco, which indicated that NtPHGPx may be involved in plant response to salt stress by affecting the scavenging activity of reactive oxygen species in tobacco.