Pcdh cmv mcs ef1 gfp puro

Short hairpin RNA (shRNA) of cTFRC were synthesized by GenePharma (Shanghai, China), shcTFRC targeting to the junction region of the cTFRC sequence. The shRNA plasmid of TFRC was purchased from Santa Cruz (Dallas, USA). EJ and T24 cells were transfected with cTFRC and TFRC shRNAs plasmids using Lipofecamine 2000 (Invitrogen, Carlsbad, CA, USA). The sequences of the effective shRNAs were provided in Additional file 1: Table S2. The full-length cTFRC cDNA was cloned into pCDH-CMV-MCS-EF1-GFP + Puro (Geneseed Biotech, Guangzhou, China) to obtain the pCDH-cTFRC overexpression of cTFRC. Production of lentiviral particles and transduction of BC cells was performed as described [14 (link)].

The ADAR1 shRNA or scramble sequence was inserted into pLKO1 vector for lentivirus generation. The ADAR1 p110 overexpression plasmid (Z3143) was purchased from GeneCopoeia (Rockville, MD, USA) and converted into pWPI vector by the Gibson assembly method (Supplementary Figure S1b. A promoter of ADAR1 p110 was obtained from genomic DNA of 293T cell by Phusion High-Fidelity DNA Polymerase (NEB, Beverly, NY, USA) and ligated into pGL3-basic vector (Promega, Madison, WI, USA) by the Gibson assembly method. For the ARE mutation, quickchange was used according to the manufacturer’s instruction. For the generation of CircARSP91 (hsa_circ_0085154) overexpression plasmid, we added 300 bp upstream and 300 bp downstream sequence to CircARSP91 nonlinear splice site (total 691 bp). Then, this fragment was inserted into pCDH-CMV-MCS-EF1-GFP+Puro (Geneseed Biotech, Guangzhou, China) to obtain the pCDH-CircARSP91.^{12 (link)} Overexpression of CircARSP91 was verified (Supplementary Figure S1C). See Supplementary Table S1 for detailed sequences.
For luciferase assays, cells were plated in 24-well plates with transfection using Lipofectamine3000 (Invitrogen) according to the manufacturer’s instruction. pRL-TK was used as the internal control. Luciferase activity was measured by Dual-Luciferase Assay (Promega) according to the manufacturer’s manual.