Construction of cDNA representational difference analysis based on two cDNA libraries and identification of garlic inducible expression genes in human gastric cancer cells

Abstract

Aim: To elucidate molecular mechanism of chemopreventive efficacies of garlic against human gastric cancer (HGC). Methods: HGC cell line BGC823 was treated with Allitridi (a kind of garlic extract) and Allitridi-treated and parental BGC823 cDNA libraries were constructed respectively by using λZAP II vector. cDNA Representational Difference Analysis (cDNA RDA) was performed using BamH I cutting-site and abundant cDNA messages provided by the libraries. Northern blot analysis was applied to identify the obtained difference products. Results: Two specific cDNA fragments were obtained and characterized to be derived from homo sapiens folate receptorα (FRα) gene and calcyclin gene respectively. Northern blot results showed a 4-fold increase in FRα gene expression level and 9-fold increase in calcyclin mRNA level in BGC823 cells after Allitridi treatment for 72 h. Conclusion: The method of cDNA RDA based on cDNA libraries combines the high specificity of cDNA RDA with abundant cDNA messages in cDNA library; this expands the application of cDNA library and increases the specificity of cDNA RDA. Up-regulation of FRα gene and calcyclin gene expressions induced by Allitridi provide valuable molecular evidence for the efficacy of garlic in treating HGC as well as other diseases.