Isolation of Genes Related to Malignant Transformation Induced by Dihydroxyepoxy Benzo Pyrene with cDNA Representational Difference Analysis

ObjectiveTo isolate the differentially expressed genes between transfo rmed cells by dihydroxyepoxy benzo pyrene and controls.MethodsmRNAs from these cells were extracted and double stranded cDNAs were prepared.Double stranded cDNAs were digested with Sau3AI,adaptor ligated,and amplified by PCR.cDNA from dihydroxy epoxy benzo pyrene-treated cells was used as a "tester",and cDNA from controls as a "driver",driving the process of subtraction.ResultsFive differential cDNA fragments were isolated by cDNA representational difference analysis(cDNA-RDA)after three rounds of subtractive hybridization.They showed five distinctive bands below 210bp on agarose gel.While these differential cDNA fragments were hybridized to total RNA by Northern Blot,it was found that they really came from the BPDE-treated cells.ConclusionscDNA-RDA was an effective,sentitive and specific technique to screen the candidates of genes related to chemical carcinogenesis.