Angermeyer, Angélica Melo; Assef, Pablo Díaz; Cifuentes, Constanza Fritz; Allaire, Carmen Gloria Artigas; Strauch, Juan Carlos Roa
December 2012
Revista Chilena de Tecnología Médica;2012, Vol. 32 Issue 2, p1713
Academic Journal
DNA methylation, especially in promoter regions of tumor suppressor genes (GST), is the primary mechanism that causes epigenetic gene silencing. The p53 gene is a major GST, responsible for regulating the expression of various genes that preserve the integrity of DNA. Objective: To standardize a protocol for the detection of methylated points in the promoter and exon 1 of the p53 gene using restriction enzymes sensitive to methylation and chain reaction of the polymerase (ER-PCR). DNA was used as a control commercial genomic DNA and DNA samples from healthy individuals. The control DNA was methylated in vitro and tested at different concentrations and digestion times varying enzyme concentration. Enzymatic Postdigestión PCR was performed. Results: incomplete enzymatic digestion was obtained with control DNA methylated and unmethylated > 0.05 ug / ul. Complete digestion was observed when using 0.05 ug / ul DNA, 10 U of enzyme and 8 hours digestion. Protocol was applied to clinical samples detected 2/15 methylated. The methodology ERSM-PCR allowed a rapid and timely analysis of methylation in the p53 gene, proving to be simple, little laborious and moderate cost.


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