PARCLIP analysis of YBX1, ELAVL1 and ALYREF to detect their binding targets

The PAR-CLIP sample was separated by SDS-PAGE, RNA were recovered from the gel by D-tubeTM Dialyzer Midi(Millipore).The mixture were digested by 4 μg/μl proteinase K (Roche, 03115828001) and precipitated in ethanol with the help of glycogen (Thermo, R0551). RNAs in ethanol were used for the small RNA library construction (NEB, E7300L). Finally, the library was sequenced on the Illumina HiSeq X-Ten platform with paired-end 150 bp (PE150) kits. Only the forward reads were used in the data analysis and uploaded. Bowtie was applied to map sequencing reads against hg19 genome with up to two mismatches allowed. PARalyzer software was used to define proteins binding clusters. Examination of YBX1, ELAVL1 and ALYREF binding targets