〔Abstract〕 To explore the clinical characteristics of smear negative pulmonary tuberculosis and the diagnostic value of multi-color nested real-time fluorescence quantitative nucleic acid amplification(Gene Xpert-MTB/RIF). The clinical data of 400 patients were analyzed retrospectively, and the value of Gene Xpert-MTB/RIF in the diagnosis of smear negative pulmonary tuberculosis was discussed. The results showed that the incidence of fever, cough/expectoration and neutrophil increase in smear negative pulmonary tuberculosis was significantly lower, the proportion of diabetes mellitus and chronic obstructive pulmonary disease was small, and the incidence of pleurisy and pulmonary fibrosis in imaging was higher than that in smear positive pulmonary tuberculosis. Gene Xpert-MTB/RIF technology has a better diagnostic value than traditional inspection methods, which is worthy of clinical promotion.