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A Protocol for Detecting DNA Methylation Changes at CpG Sites of Stemness-Related Genes in Aging Stem Cells
Mahshid Hodjat1, Seyed Mojtaba Daghighi2, Mohammad Abdollahi3,4
1Dental Research Center, Dentistry Research Institute, Tehran University of Medical Sciences, Tehran, Iran.
Abstract:
Aging adversely affects the self-renewal and differentiation capabilities of stem cells, which impairs tissue regeneration as well as the homeostasis. Epigenetic mechanisms, specifically DNA methylation, play a key role in the maintenance of pluripotency in stem cells and regulation of pluripotency-related gene expression. Age-related modifications in methylation patterns could influence the expression of genes critical for stem cell potency maintenance, including transcription factors Nanog and Sox2. The following chapter describes a step-by-step bisulfite sequencing protocol for detection of methylation changes in the aging stem cells and provides valuable insights into the stem cells epigenetic profile. Further, the methodology describes the steps of genomic DNA extraction, bisulfite conversion, real-time PCR amplification, and sequencing for an in-depth view of the epigenetic profile derived from aging stem cells.
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