Labeling and sequencing nucleic acid modifications using bio-orthogonal tools

Abstract

The bio-orthogonal reaction is a type of reaction that can occur within a cell without interfering with the active components of the cell. Bio-orthogonal reaction techniques have been used to label and track the synthesis, metabolism, and interactions of distinct biomacromolecules in cells. Thus, it is a handy tool for analyzing biological macromolecules within cells. Nucleic acid modifications are widely distributed in DNA and RNA in cells and play a critical role in regulating physiological and pathological cellular activities. Utilizing bio-orthogonal tools to study modified bases is a critical and worthwhile research direction. The development of bio-orthogonal reactions focusing on nucleic acid modifications has enabled the mapping of nucleic acid modifications in DNA and RNA. This review discusses the recent advances in bio-orthogonal labeling and sequencing nucleic acid modifications in DNA and RNA.

Fig. 1. A schematic diagram of the simplified process for labeling modified bases using bio-orthogonal tools.

Fig. 2. The transition of various cytosine modifications.

Fig. 3. The chemical method for labeling DNA modifications.

Fig. 4. The chemical method for labeling RNA modifications.

Fig. 5. Enzyme-assisted methods for labeling nucleic acid modifications.

Fig. 6. Metabolic labeling by nucleoside analogs. (A) The chemical structures of nucleoside analogs. (B) Metabolic labeling of RNA. (C) Cell-specific metabolic labeling.

Fig. 7. Mapping nucleic acid modifications based on bio-orthogonal tools. (A) The workflow of mapping. (B) Mapping 5hmC's position in the genome.

Fig. 8. Base resolution sequencing methods for nucleic acid modifications. (A) fC-CET. (B) hmC-CATCH. (C) Click-code-seq for labeling 8-oxoG. (D) caCLEAR. (E) CeU-Seq.

Hui Liu

Yafen Wang

Xiang Zhou