Pierce magnetic pull down kit

The full‐length sense and antisense sequences of the FASRL fragment were cloned into the pcDNA3.1 vector using the HindIII restriction enzyme (Thermo Fisher Scientific, FD0504) and XhoI restriction enzyme (Thermo Fisher Scientific, FD0694). A TranscriptAid T7 High Yield Transcription Kit (Thermo Fisher Scientific, K0441) was used to transcribe the full‐length sense and antisense RNA sequences of FASRL in vitro, and then, these sequences were purified using a GeneJET RNA Purification Kit (Thermo Fisher Scientific, K0731). A Pierce RNA 3′ End Desthiobiotinylation Kit (Thermo Fisher Scientific, 20163) was used to label the purified sense and antisense RNA sequences of FASRL with desthiobiotin at the 3′ end. Next, the lncRNA–protein complexes were purified by a Pierce Magnetic Pull‐Down Kit (Thermo Fisher Scientific, 20164) and separated by SDS‐PAGE. The separated proteins were stained with a Fast Silver Stain Kit (Beyotime, P0017S), and then, MS was performed to identify the specific proteins that bound to the FASRL. Finally, immunoblotting was used to verify that the proteins interacted with FASRL.

The interaction between circACAP2 and miR-193a-5p was measured by a Pierce™ Magnetic Pull-Down Kit (Thermo, USA) following the manufacturer's protocol. The biotin-labeled miR-193a-5p probe and the negative control were synthesized by RiboBio (China).