Eight complementary DNA (cDNA) clones highly expressed in fetal rat stomach but not in normal adult rat stomach were isolated after screening 2 x 10(4) independent recombinants from a subtracted cDNA library. The cDNA library was first prepared from RNAs of total stomach at 16 days gestational period, and this cDNA library was subtracted by cDNAs prepared from adult rat total stomach RNA, using a novel PCR-based cDNA subtraction method. Northern blot analysis revealed that as many as six of eight clones thus isolated were overexpressed in at least some of the human or rat gastric cancers. From analysis of partial nucleotide sequence, four cDNA clones were identified as profilin, pro-alpha 1 (1) collagen, nucleolar protein B23.2, and elongation factor 1 alpha subunit. The remaining two clones were derived from novel genes. These novel genes, L-1 and L-2, are developmentally well regulated in the stomach. The present results clearly show that genes expressed preferentially in embryo stomach are most likely to be highly expressed in gastric cancer. The method described here provides us with a rapid method for identification of genes with significantly increased expression in cancer.