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Optimization of MethylRAD sequencing to detect changes in DNA methylation at enhancer elements in differentiating embryonic stem cells

ABSTRACT: Optimization of MethylRAD sequencing to detect changes in DNA methylation at enhancer elements in differentiating embryonic stem cells

PROVIDER: PRJNA665721 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR12713663.fastq.gz	Fastqsanger.gz
	SRR12713664.fastq.gz	Fastqsanger.gz
	SRR12713665.fastq.gz	Fastqsanger.gz

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Optimization of MethylRAD sequencing to detect changes in DNA methylation at enhancer elements in differentiating embryonic stem cells

Project description:Differential DNA methylation is characteristic of gene regulatory regions, such as enhancers, which mostly constitute low or intermediate CpG content in their DNA sequence. Consequently, quantification of changes in DNA methylation at these sites is challenging. Given DNA methylation across most of the mammalian genome is maintained, use of genome-wide bisulfite sequencing to measure fractional changes in DNA methylation at specific sites is an overexertion which is both expensive and cumbersome. Here we developed a MethylRAD technique with an improved experimental plan and bioinformatic analysis tool to examine regional DNA methylation changes in embryonic stem cells (ESCs) during differentiation. The transcriptional silencing of pluripotency genes (PpGs) during ESC differentiation is accompanied by PpG enhancer (PpGe) silencing mediated by demethylation of H3K4me1 by LSD1. Our MethylRAD data show that in presence of LSD1 inhibitor, a significant fraction of LSD1-bound PpGe fails to gain DNA methylation. We further show that this effect is mostly observed at PpGe with low/intermediate CpG content. Underscoring the sensitivity and accuracy of MethylRAD sequencing, our study demonstrates that this method can detect small changes in DNA methylation at regulatory regions including with low/intermediate CpG content, thus asserting it use as a method of choice for diagnostic purpose.

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