GSM4942752: Empty vector FLAG IP 2; Homo sapiens; OTHER
Cells were lysed with RNase I to fragment RNA as previously described (Van Nostrand, E.L. et al. (2017) Methods Mol Biol). FLAG antibody (F-1084, Sigma-Aldrich) was then pre-coupled to Protein G Dynabeads (Thermo Fisher), added to lysate, and incubated overnight at 4 degrees Celcius. Prior to immunoprecipitation, 2% of the sample was taken as the paired input sample, with the remainder magnetically separated and washed with lysis buffer only. Library construction was performed by excising the area from ∼65 kDa to ∼140 kDa. RNA adapter ligation, IP-western, reverse transcription, DNA adapter ligation, and PCR amplification were performed as previously described (Van Nostrand, E.L. et al. (2017) Methods Mol Biol). Single-end Enhanced Crosslinking and Immunoprecipitation (seCLIP) by Eclipse Bioinnovations Inc (San Diego, www.eclipsebio.com) according to the published protocol (Van Nostrand, E.L. et al. (2017) Methods Mol Biol).
GEO Accession
GSM4942752