Abstract
DNA methylation is a fundamental epigenetic process and have a critical role in many biological processes. The study of DNA methylation at a large scale of genomic levels is widely conducted by several techniques that are next-generation sequencing (NGS)-based methods. Methylome data revealed by DNA methylation next-generation sequencing (mNGS), should be always verified by another technique which they usually have a high cost. In this study, we offered a low-cost approach to corroborate the mNGS data. In this regard, mNGS was performed on 6 colorectal cancer (case group) and 6 healthy individual colon tissue (control group) samples. An R-script detected differentially methylated regions (DMRs), was further validated by high resolution melting (MS-HRM) analysis. After analyzing the data, the algorithm found 194 DMRs. Two locations with the highest level of methylation difference were verified by MS-HRM, which their results were in accordance with the mNGS. Therefore, in the present study, we suggested MS-HRM as a simple, accurate and low-cost method, useful for confirming methylation sequencing results.
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Acknowledgements
Our sincere thanks go to Mr. Ebrahim Pouladin and Mrs. Nafiseh Shalchi for their close support in colorectal cancer research programs. We would also give special thanks to Dr. Abdorasoul Hayatbakhsh and Dr Ghodratollah Soltani for their cooperation in this study.
Funding
This study was supported financially by Reza Radiotherapy and Oncology Center, Mashhad University of Medical Sciences (Grant number: 961906) and Iran National Science Foundation (INSF; proposal number: 93048371), Iran.
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This study was approved by ethics committee of Mashhad University of Medical Sciences, Mashhad, Iran (ethic code: IR.MUMS.MEDICAL.REC.1399.642).
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Communicated by Shuhua Xu.
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438_2022_1906_MOESM1_ESM.png
Supplementary file1 Methylation Call (MC) for case and control samples using primer A and B sets. The size of the symbols correspond to coverage (PNG 252 KB)
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Javadmanesh, A., Mojtabanezhad Shariatpanahi, A., Shams Davodly, E. et al. MS-HRM protocol: a simple and low-cost approach for technical validation of next-generation methylation sequencing data. Mol Genet Genomics 297, 1101–1109 (2022). https://doi.org/10.1007/s00438-022-01906-1
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DOI: https://doi.org/10.1007/s00438-022-01906-1