Identification of Target LncRNAs of IMP1 in T47D Cells by RIP-Seq technique

IMP1 is a RNA binding protein shown to regulate translation, localization, or stability of their target RNAs. However, whether IMP1 can widely bind to lncRNAs and regulate their expression in Breast cancer cells remains to be confirmed. In this study, we used RNA binding protein immunoprecipitation-high throughput sequencing (RIP-Seq) technique to identify LncRNAs which bind to IMP1 protein in breast cancer cell line T47D cells. Cell lysates were prepared from cultured T47D cells and then incubated with protein A beads coated with anti-IMP1 antibody or normal IgG control at 4°C for 4 h in the presence of RNasin (200 U/mL) and protease inhibitors. After incubation, the supernatant was removed by a short centrifugation and the beads were washed three times in ice-cold lysis buffer. Immunoprecipitates were first verified by western blotting using the anti-IMP1 antibody. Total RNAs was extracted from individual precipitates with TRIzol reagent. The binding LncRNAs were analyzed by LncRNA-seq. The RNA-Seq experiments and data analysis were performed at the Shanghai Biotechnology Corporation in Shanghai, China.