Objective: To search for new putative tumor suppressor genes in colorectal carcinoma.
Methods: Subtractive hybridization technologies were applied to screen and select genes, the expression of which was down-regulated in colorectal carcinoma. mRNAs uniquely expressed in normal cells but not in colorectal carcinoma were recovered as cDNA (sub-cDNA) after two rounds of subtractive hybridization with mRNA prepared from colorectal carcinoma. The sub-cDNAs were then used as probes to screen a normal human colon cDNA library constructed in lambda-Zap II phage. The DNAs of positive clones were in vivo excised, and partial DNA sequences were analyzed and compared with DNA sequence database Genbank.
Results: A total of 46 different clones with an average of about 1 kilobases in transcript size was recovered. Among these 46 down-regulated genes in colorectal carcinoma were genes encoding immunoglobulin (n = 32), 40-kDa keratin intermediate filamentous protein or IFP (n = 1), major histocompatibility complex-related protein (n = 1), unrelated structural proteins (n = 10) and gene products yet to be identified (n = 2). RNA dotblot hybridizations confirmed that all 46 clones contained genes that were down-regulated and have not been reported before in colorectal carcinoma.
Conclusion: The results of this study suggested that the 46 clones were down-regulated in colorectal carcinoma, they should be further studied as new putative tumor suppressor genes and could be used as new tumor markers of colorectal carcinoma.