title: SRSF1 RIP-Seq of Homo sapiens: LCLs after ionizing radiation
design: Reverse immunoprecipitation was carried out using Magna RNA-Binding Protein Immunoprecipitation Kit (Millipore) following the manufacturers protocol. Briefly, for each immunoprecipitation reaction, 2  10^7 cultured human B-cells from GM11994 and GM12878 cell lines were harvested in three conditions (no irradiation, and two hours post irradiation, and six hours post irradiation). Cells were lysed in 100 L lysis Buffer with protease and RNase inhibitors. Ten micrograms of anti-SRSF1 antibody (Bethyl, #A302-052A) and negative control rabbit IgG (Millipore, #12-370) were conjugated to Magnetic Protein A/G beads. One hundred microliters of cell lysate was added into 900 L Immunoprecipitation Buffer with RNase inhibitor and incubated with 50 L beads-antibody complex overnight at 4C. Bead-bound immunoprecipitates were then washed six times using cold Wash Buffer with RNase inhibitor andincubated with protease K in the presence of 1% SDS for 30 min at 55C. RNA and DNA were then extracted from supernatants using phenol:chloroform:isoamyl alcohol and precipitated using ethanol. Precipitated RNA was digested by DNase I (DNA-free kit, Ambion). RNA-seq libraries for SRSF1 immunoprecipitate and input RNA were prepared as described above. Sequencing reads were preprocessed and aligned as described above.