Detection of p16 Promoter Hypermethylation by Methylation-Specific PCR

Abstract

DNA methylation plays a decisive role in the regulation and control of gene expression. DNA methylation is a covalent modification, in which a methyl group is attached to the 5th carbon of the cytosine ring of a CpG dinucleotide that is located upstream from the promoter region of a gene. Promoter hypermethylation (gain of DNA methylation) of the p16 gene may cause silencing of gene expression and plays an important role in cancer. Therefore, detection of the methylation status of p16 gene is an important tool in epigenetic studies of various human cancers. The methylation-specific PCR (MSP) is the most commonly used technique for studying DNA methylation. This technique is based on bisulfite modification of DNA, which converts unmethylated cytosine (C) into uracil (U) and leaving methylated cytosine (C) unchanged. Here we describe the bisulfite modification of DNA samples and detection of promoter methylation of p16 gene from bisulfite-treated DNA using MSP. In MSP, modified DNA samples are subjected to PCR amplification using methylated and unmethylated specific primers for the p16 gene separately. The PCR amplified products are then analyzed in a 2.5-3% agarose gel containing ethidium bromide. The PCR amplified band generated by specific sets of primers is used to determine the methylation status of the p16 gene.

DNA methylation detection methods used in colorectal cancer

“…As an endpoint analysis technique[40], the primary limitation of MSP is that it yields qualitative results, meaning that well-standardized MSP assays provide information that is restricted to the presence of methylated, unmethylated, or both methylated and unmethylated alleles[41].…”

GABRP is a potential prognostic biomarker and correlated with immune infiltration and tumor microenvironment in pancreatic cancer

“…We further explored the potential mechanism of GABRP overexpression in pancreatic cancer from the perspective of epigenetics. DNA methylation plays an important role in regulating gene expression and aberrant DNA methylation is involved in cancer progression ( 45 ). DNA hypomethylation is frequently observed in human tumors and contributes to the aberrant activation of specific genes related to cancer progression ( 46 ).…”

Developments and future prospects of personalized medicine in head and neck squamous cell carcinoma diagnoses and treatments

“…The detection of p16 gene hypermethylation is often used as a biomarker for the early diagnosis and prognosis of HNSCC (Figure 2). Methylation‐specific polymerase chain reaction (MSP) and other molecular techniques are commonly employed to assess the methylation status of the p16 gene in tumor samples 42 . The presence of p16 hypermethylation may indicate an increased risk of HNSCC, and also it can act as a potential therapeutic target 38 .…”

“…Methylation‐specific polymerase chain reaction (MSP) and other molecular techniques are commonly employed to assess the methylation status of the p16 gene in tumor samples. 42 The presence of p16 hypermethylation may indicate an increased risk of HNSCC, and also it can act as a potential therapeutic target. 38 Research has shown that the methylation status of the DAPK gene promoter is altered in HNSCC and is associated to its progression, prognosis, and response to therapy.…”