Expression profiling by high throughput sequencing Methylation profiling by high throughput sequencing
Summary
Investigations of 5-hydroxymethylcytosine (5hmC) in biologically and clinically samples and models with low cell numbers have been hampered by the low sensitivity and reproducibility using current 5hmC mapping approaches. Here, we develop a selective 5hmC chemical labeling approach using tagmentation-based library preparation in order to profile nanogram levels of 5hmC isolated from ~1,000 cells (nano-hmC-Seal). Using this technology, we profiled the dynamics of 5hmC across different stages of mouse hematopoietic differentiation. Additionally, applying nano-hmC-Seal to the hematopoietic multipotent progenitor cells in an acute myeloid leukemia (AML) mouse model, we identified leukemia-specific, differentially hydroxymethylated regions that harbor previously reported and as-yet-unidentified functionally relevant factors. The change of 5hmC patterns in AML strongly correlates with the altered gene expression on a global scale. Together, our new approach offers a highly sensitive and robust method to study and detect DNA methylation dynamics from in vivo model and clinical samples.
Overall design
Selective 5hmC chemical labeling approach using tagmentation-based library preparation in order to profile nanogram levels of 5hmC isolated from ~1,000 cells
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