Objective: To observe the changes of the concomitant gene mutation/amplification in patients with lung cancer based on next generation sequencing (NGS) technology and analyze its clinical significance. Methods: Retrospective analysis was performed on 71 lung cancer patients (132 samples) detected by NGS technology in the Second Affiliated Hospital of Soochow University from December 2015 to July 2020, including 67 blood samples, 48 tumor tissues, 15 pleural effusion and 2 cerebrospinal fluid. GraphPad prism7.0 statistical software was used for statistical analysis of concomitant gene mutation/amplification in different clinical samples. Results: In addition to EGFR mutation, mutations/amplification of the first 5 genes were: TP53, KRAS, ERBB2, RB1 and MET. A total of 50 newly diagnosed patients' blood samples were detected by NGS technology. The results showed that EGFR positive rate was 34% (17/50). Except EGFR mutation, there were 79 kinds of gene mutations/amplifications detected, with an average of 2.46 times/example. But with no statistical difference in concomitant gene mutation/amplification among clinical stage, smoking status, age, gender and pathological classification groups in blood samples. A total of 46 newly diagnosed patients' tissue samples were detected by NGS technology. Of which EGFR positive rate was 50% (23/46). Except EGFR mutation, there were 160 kinds of gene mutations/amplifications detected, with an average of 6.20 times/example. There was no statistical difference in concomitant gene mutation/amplification between clinical stage and age groups in tissue samples（P＞0.05）. But in terms of smoking status, gender and pathological classification groups, we could find statistically difference (U=74.000，P＜0.001；U=130.5，P=0.003；F=8.968，P=0.011）. At the same time, TP53 mutation or not had no statistical significance on percent survival（the blood group: x2=0.321，P=0.571; the tissue group：x2=0.309，P=0.579）. For the same patient, the frequency of gene mutation/amplification in the tissue group（[3（1，8）]）was higher than that in the blood group [3（0，1）] of the same patient, and the difference was statistically significant (W=-150，P =0.001). In the 17 patients with dynamic monitoring, there was no statistically significant difference in the frequency of concomitant gene mutation/amplification between the two groups before and after anti-tumor treatment(W=-3，P=0.916). Conclusion: There may be multiple parallel gene mutations/amplifications before the first-line treatment for lung cancer. Previous smoking and male patients were associated with higher levels of mutation/amplification status. Changes in the frequency of gene mutation/amplification cannot indicate disease progression, and TP53 mutation has no significant effect on the percent survival of patients.