Construction Of A Stubtracted CDNA Library And Cloning Of Differentially Expressed Genes In Stomach Signet Ring Cell Carcinoma

Objectives:To isolate genes expressed exclusively or preferentially in human stomach signet ring cell carcinoma in an effort to explain the mechanisms of the tumor.Methods: Suppression subtractive hybridization (SSH) was performed to isolate up-regulated cDNA fragments in signet ring cell carcinoma of the stomach tissue. Human gastric signet ring cell carcinoma tissue was used as the tester, and adjacent normal stomach tissue was used as the driver. Then these cDNA fragments were directly inserted into T/A cloning vector to set up a subtraction cDNA library of specially or highly expressed genes in gastric signet ring cell carcinoma . Some clones were randomly picked and identified to have contained insert sizes ranging from 150 to 1000bp. some of these clones were sequenced and compared with known sequences in the public databases of GenBank/EMBL/DDBJ using NCBI BLAST. The functions of these genes on the development of tumor have been analysis.Results: 1. A subtracted cDNA library of differentially expressed genes in human gastric signet ring cell carcinoma is constructed successfully with SSH and T/A cloning techniques.2. Some differentially expressed gene fragments have been cloned: one was similar to chromosome sequence and has been submitted to ESTdatabase (dbEST) at NCBI as an EST of a new given gene(Accn:CN446913),the others were from previously described genes,a large proportion of witch had been reported to be linked with tumor . Conclusion:1. SSH is an effective way to isolate differentially expressed genes. 2. Genes expressed differentially may have played an important role in the development and growth of human gastric signet ring cell carcinoma .3. An EST of a new given gene was obtained and might be useful in revealing the mechanisms of gastric signet ring cell carcinoma.