Protocol detail

Quantitative Methylation-Specific PCR Analysis

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Aberrant DNA methylation, which often occurs around the transcriptional start site (TSS) within a CpG island, was evaluated using Q-MSP. The sequences of primers used in this study are shown in Supplementary Table S3. A standard curve for Q-MSP was constructed by plotting five serially diluted standard solutions of EpiScope methylated HeLa gDNA (TaKaRa). To analyse the normalised methylation value (NMV) of the PCR conditions, an analytical method was used as previously described [44 (link)].

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Misawa K., Imai A., Matsui H., Kanai A., Misawa Y., Mochizuki D., Mima M., Yamada S., Kurokawa T., Nakagawa T, & Mineta H. (2020). Identification of novel methylation markers in HPV-associated oropharyngeal cancer: genome-wide discovery, tissue verification and validation testing in ctDNA. Oncogene, 39(24), 4741-4755.

Publication 2020

EpiScope methylated HeLa gDNA

Cpg island Dna methylation Hela Methylation Primers Transcriptional start site

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Variable analysis

independent variables

Aberrant DNA methylation

dependent variables

Normalised methylation value (NMV)

control variables

Sequences of primers used in the study (provided in Supplementary Table S3)
Standard curve for Q-MSP constructed using serially diluted standard solutions of EpiScope methylated HeLa gDNA (TaKaRa)

controls

Positive control: EpiScope methylated HeLa gDNA (TaKaRa)
Negative control: Not explicitly mentioned

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