A cross-linking approach to map small molecule-RNA binding sites in cells.

ABSTRACT: Methods to identify RNAs bound by small molecules in cells are sparse. Herein, an advance to identify the direct RNA targets of small molecules in cells is described. The approach, dubbed Chemical Cross-Linking and Isolation by Pull-down to Map Small Molecule-RNA Binding Sites (Chem-CLIP-Map-Seq), appends a cross-linker and a purification tag onto a small molecule. In cells, the compound binds to RNA and undergoes a proximity-based reaction. The cross-linked RNA is purified and then amplified using a universal reverse transcription (RT) primer and gene-specific PCR primers. At nucleotides proximal to the binding site, RT "stops" are observed. This approach has broad utility in identifying and validating the RNA targets and binding sites of small molecules in the context of a complex cellular system.

SUBMITTER: Velagapudi SP 

PROVIDER: S-EPMC6598432 | biostudies-literature | 2019 Jun

REPOSITORIES: biostudies-literature