Objective The objective of this study is to investigate the methylation status of zonula occluden-1 (ZO-1) gene in patients with non-small cell lung cancer (NSCLC) and to indentify this roles in pathogenesis,development and classification of NSCLC.Methods The methylation status of ZO-1 gene of 101 patients with NSCLC and 61 patients with benign lung disease were detected by methylationspecific-polymerase chain reaction (MS-PCR). ZO-1 mRNA and protein of 200 unmethylation tissue and 62 methylation tissue were detected by real-time PCR and Western blotting separately. Result Both of ZO-1 mRNA and protein was significant statistic different between methylation group and unmethylation group(EM>t/EM>=-26.028,P<0.01,EM>t/EM> =-37.216,EM>P/EM><0.01).There was significant statistic difference between carcinoma tissue group(32.7%,33/100) and control group(11.5%,7/61) （EM>χ/EM>SUP>2/SUP>=9.190,EM>P/EM><0.01）. ZO-1 promoter methylation status in adjacent tissues was significant statistic different between lymph node invasion group(35.5%,11/31) and lymph nodes without invasion group(17.1%,12/70) （EM>χ/EM>SUP>2/SUP>=4.110,EM>P/EM><0.05）. ZO-1 promoter methylation status of carcinoma tissues wase significant statistic different between 2-year suvival group (27.5%,14/51) and 2-year death group(43.2%,19/44) (EM>χ/EM>SUP>2/SUP>=4.150, EM>P/EM><0.05）. Conclusion Methylation status of ZO-1 promoter may affect the transcription of mRNA and expression of protein. ZO-1 promoter methylation status of adjacent tissues and carcinoma tissues in N