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Unified analysis of DNA methylation, hydroxymethylation and chromatin accessibility through targeted DNA labeling [TOP-Seq]

ABSTRACT: Unified analysis of DNA methylation, hydroxymethylation and chromatin accessibility through targeted DNA labeling [TOP-Seq]

PROVIDER: PRJNA970253 | ENA |

REPOSITORIES: ENA

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Unified analysis of DNA methylation, hydroxymethylation and chromatin accessibility through targeted DNA labeling

Project description:Unified analysis of DNA methylation, hydroxymethylation and chromatin accessibility through targeted DNA labeling

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Unified analysis of DNA methylation, hydroxymethylation and chromatin accessibility through targeted DNA labeling [TOP-Seq]

Project description:Epigenetic regulation depends on complex interactions between different epigenetic factors, such as DNA methylation and chromatin accessibility. As the gene activation potential cannot be predicted by looking at chromatin accessibility or DNA modification alone, methods capable of relating multiple factors are highly desirable. Existing methods for combined chromatin accessibility and DNA modification analysis rely on bisulfite sequencing, which has a number of well known drawbacks. We developed a novel cost-efficient method for simultaneous profiling of DNA methylation and hydroxymethylation within open chromatin loci. Our approach is based on chemo-enzymatic covalent tagging of unmodified (uCG) and hydroxymethylated (5hmC) CG sites along with GC sites in native chromatin, which are then mapped by tag-selective sequencing. Validation of the technology on model DNA systems and mammalian cells demonstrated a great applicability of this method for epigenomic studies. We employed the technology for tracking chromatin dynamics combined with changes of two different cytosine states, unmodified and hydroxymethylated, during differentiation of embryonic stem cells (ESC) which revealed extensive chromatin dynamics and DNA modification changes across gene bodies of development-associated genes at early transition from ESC. Strong DNA demethylation in the ESC-to-neuron differentiation pathway proceeds independently from cell transcriptional programs and highlights the value of separate 5hmC and 5mC analysis in understanding cell fate decisions.

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Project description:This SuperSeries is composed of the SubSeries listed below.

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Project description:In this study we addressed the question whether changes of DNA hydroxymethylation after X-ray irradiation occur in a random fashion or if they show preferences for specific regions or features. We did not only focus on gene loci and promotor sights but also on gene body hydroxymethylation alterations and associated changes in gene expression. A more profound insight into hydroxymethylation characteristics triggered by radiation could provide clues to mechanisms and consequences of cellular response to irradiation including the immediate consequence of DNA strand breaks associated with active DNA demethylation as well as the long term risk of tumorigenesis or other associated diseases.

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Project description:Archaeological materials are a finite resource, and efforts should be made to minimize destructive analyses. This can be achieved by using protocols combining extraction of several lines of evidence, which decreases the material needed for analyses while maximizing the information yield. Archaeological dental calculus is a source of several different types of biomolecules, as well as microfossils, and can tell us about the human host, microbiome, diet, and even occupational activities. Here, we present a unified protocol allowing for simultaneous extraction of DNA and proteins from archaeological dental calculus. We evaluate the protocol on dental calculus from a range of ages and estimated preservation states, and compare it against standard DNA-only and protein-only protocols. We find that most aspects of downstream analyses are unaffected by the unified protocol, although minor shifts in the recovered proteome can be detected. Protein recovery depends on both the amount of starting material and choice of extraction protocol, whereas DNA recovery is significantly lowered through the unified protocol. However, DNA recovery from dental calculus is generally very high, and we found no differences in DNA fragment characteristics or taxonomic profile. In conclusion, the unified protocol allows for simultaneous extraction of two complementary lines of evidence from archaeological dental calculus without compromising downstream analyses, thereby minimizing the need for destructive analysis of this finite resource.

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