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2005
LSU-HHMI Summer Undergraduate Research Program |
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Eric Harris (Troy) and Vince Wilson, Biological Sciences
Methylation-specific PCR Analyses of DNA from Norma, Neoplastic, and Cancerous Skin Specimens
The addition of methyl-groups to cytosines residues in CpG islands is involved in the transcription inactivation of two tumor suppressor genes (p16 and DAPK). The p16 gene protein product is an inhibitor of cell division. The DAPK gene (death-associated protein kinase) codes for a protein involved in apoptosis (programmed cell death). The absence of either or both of these genes can lead to the uncontrolled cell division characteristic of neoplasia. Analyzing, cataloging, and understanding the patterns of the CpG islands of these genes is important for understanding tumor suppressor gene silencing in human cancer. MSP (methylation-specific PCR) can be used to determine the methylation status of these CpG sites. MSP requires that the template DNA be first modified with a sodium bisulfite treatment, which converts all unmethylated, but not methylated, cytosines to uracil. Methylated- and unmethylated-specific primers are then used to amplify the DNA. In this study, we demonstrate the use of MSP to identify hypermethylation changes in the promoter regions of p16 and DAPK in DNA extracted from normal, pre-neoplastic, and squamous cell carcinoma skin specimens of female vulvar tissue.
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