Few target molecules have been identified that enable the diagnosis of lung cancer with high sensitivity and specificity, especially in the early clinical stages. Herein, we present the first evidence for mRNA overexpression of SALL4, a transcription factor essential for embryonic development and the self-renewal of embryonic stem cells, in lung cancer. Analysis using cancerous and noncancerous tissues revealed that the sensitivity and specificity of SALL4 mRNA were 85.1 and 92.9%, respectively, estimated using the cutoff value obtained from analyzing the receiver operating characteristic curve. Furthermore, comparison of paired tissues from the same patient revealed elevated SALL4 mRNA levels that were greater than two-fold in 93% of the specimens. SALL4 mRNA was highly expressed even in the early clinical stages and there was no difference in the positivity rate between stage IA and other stages. An siRNA approach to determine the significance of SALL4 expression revealed catastrophic growth inhibition of SBC-1 lung cancer cells that was induced by cell cycle arrest at the G1/early S phase. Therefore, SALL4 mRNA may be a candidate for use as support in the diagnosis of lung cancer, and may also represent a therapeutic target.