Sequencing uracil in DNA at single-nucleotide resolution
Jiang, L.; Yin, J.; Qian, M.; Rong, S.; Chen, K.; Zhao, C.; Tan, Y.; Guo, J.; Chen, H.; Gao, S.; Liu, T.; Liu, Y.; Shen, B.; Yang, J.; Zhang, Y.; Meng, F.-L.; Hu, J.; Ma, H.; Chen, Y.-H.
Show abstract
As an aberrant base in DNA, uracil is generated by dUMP misincorporation or cytosine deamination, and involved in multiple physiological and pathological processes. Current methods for whole-genome mapping of uracil all rely on uracil-DNA N-glycosylase (UNG) and are limited in resolution or specificity. Here, we present a UNG-independent Single-Nucleotide resolution Uracil Sequencing (SNU-seq) method utilizing the UdgX protein which specifically excises the uracil and forms a covalent bond with the resulting deoxyribose. SNU-seq was validated on synthetic DNA and applied to mammalian genomes. We found that the uracil content of pemetrexed-treated cells fluctuated along with DNA replication timing. We also accurately detected uracil introduced through cytosine deamination by the cytosine base editor (nCas9-APOBEC) and verified uracil occurrence in "WRC" motif within Activation-Induced Cytidine Deaminase (AID) hotspot regions in CSR-activated UNG -/- B cells.
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