Rnascope kit

RNAscope kit (Invitrogen, CA, United States) was used to perform RNA FISH according to manufacturer’s instructions. Briefly, the kidney tissues were fixed with 4% paraformaldehyde and embedded in paraffin. 5 μm sections were obtained by a microtome. After deparaffinization and hydration, the tissue sections were hybridized with miR-543 probe with Cy3 fluorescent dye (RiboBio Co. Ltd., Guangzhou, China) at 50°C in hybridization buffer for 3 hours, and the section was mounted onto a slide using the mounting media containing DAPI (Vector Lab, Inc., Burlingame, CA, United States). Fluorescent images were captured by Leica AM6000 microscope.

RNAScope kit (Invitrogen; Thermo Fisher Scientific, Inc.) was used to perform FISH according to the manufacturer’s instructions. Briefly, cells cultured on a coverslip were fixed with 4% paraformaldehyde and permeabilized with 0.05% Triton-X100 for 15 min. Next, 50 nM circ_0001789 probe with cyanine 3 fluorescent dye (Guangzhou RiboBio Co., Ltd.) in hybridization buffer was applied for 3 h incubation at 50 °C. Upon washing with TBST buffer, mounting medium containing DAPI (Vector Laboratories, Inc.) was used to mount the cell samples on the slides. The expression intensity and localization of circ_0001789 was observed with a fluorescence microscope.