Isolation Of The Genes Related To Breast Cancer By Suppression Subtractive Hybridization

Breast cancer is one of the most common form of malignant tumor. Many genes are involved in oncogenesis and development of breast cancer, and the erbB2 gene is one of these. Overexpression of erbB2 has been reported in numerous cancers and occurs in 30% of breast cancers, which is correlated with reduced survival. However, the biological function of erbB2 signal transduction pathways remains unclear. To identify novel genes associated with erbB2-positive mammary tumor, we constructed subtracted cDNA libraries of mouse mammary tumor tissue and normal mammary tissue using two-directional (forward and backward) suppression subtractive hybridization (SSH). Then, using T/A cloning technique, cDNA fragments of differentially expressed genes in tumor or normal mammary tissues were inserted into E. coli DH5α. We randomly picked up 492 clones and identified by colony PCR method. After differential screening by reverse dot blotting, 80 positive clones were further analyzed by DNA sequencing, GenBank and EST database searching. Sequencing results showed that 40 clones including known and novel gene/ESTs differentially expressed in mammary tumor tissue. These genes associate with transduction, cell proliferation/growth /apoptosis, cytoskeleton, membrane proteins, metabolism, secreted/ECM proteins and so on.The subtracted cDNA libraries of mouse mammary tumor tissue and normal mammary tissue were made successfully. We obtained not only some known genes but also some novel genes related to breast cancer. The known genes we obtained validated the previous research results while the novel genes laid foundation for analyzing the new genes' functions. These materials may provide abasis for further diagnosis and therapy of breast cancer.