Mucin glycoprotein can promote tumor-cell invasion metastasis and modulate the immune recognition of cancer. This study aimed to elucidate the clinical significance of mucin gene overexpression in lung cancer. We collected 60 lung cancer samples and paired non-tumorous lung portions of varying types and stages. Slot-blot analysis with specific anti-sense oligonucleotide probes derived from tandem repeat sequence of MUC1, -2, -3, -4, 5B and 5AC were utilized to compare the amount of mucin gene mRNA in tumor samples with that of the non-tumorous counterparts. A ratio higher than 1.5 for each specific mucin mRNA amount was considered to indicate mucin gene overexpression in tumors. Immunohistochemical staining of monoclonal antibodies against mature airway mucin (17Q2) and MUC1 mucin protein (HMFG2) were also used to analyze mucin protein. The study showed that overexpression of mucin genes frequently occurred in lung cancer (25 out of 60, 41.7%), but that there was no preferential expression of a particular mucin gene or a combination of mucin genes in these tumors. The overexpression of mucin genes and mucin protein had no correlation with tumor stage, nodal stage, histology or pathological differentiation grade. Tumors of smokers had higher MUC5B and MUC5AC mRNA expression ratios than those of non-smokers. Tumors with increased expression of mucin genes tended to be associated with post-operative relapse, especially when MUC5B and MUC5AC genes were overexpressed (p = 0.015 and 0.025, respectively). The study suggests that overexpression of novel tracheobronchial mucin genes may result in an increased likelihood of post-operative lung-cancer recurrence or metastases.