GSM5951892: RNMT 1 size matched input; Mus musculus; RIP-Seq
Lysis steps for seCLIP were performed as described previously (Van Nostrand et al, 2017). Samples were digested with optimised high (1:20) and low (1:200) concentrations of RNase1 as indicated. Lysates were precleared with protein G dynabeads (Thermo Fisher Scientific) at 4°C for 1hour, prior to immunoprecipitation with 50µl of protein G dynabeads preincubated with 5 µg of either RNMT, or isotype control antibody overnight at 4°C. 10% of each sample was subsequently removed to serve as the size-matched input control. IP samples were were washed three times for 5 minutes with high salt wash buffer supplemented with 2M urea. Downstream library preparation was performed as described previously (Van Nostrand et al, 2017). Libraries underwent sequencing using the Illumina MiSeq v3 150 flow cell (2x75bp reads).