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RNA immunoprecipitation (RIP)

YD Yuqing Duan
YJ Yunlong Jia
JW Jiali Wang
TL Tianxu Liu
ZC Zishuo Cheng
MS Meixiang Sang
Wl Wei lv
JQ Jing Qin
LL Lihua Liu
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The EZ-Magna RIP kit (Millipore Corp, Billerica, USA) was used for RNA immunoprecipitation (RIP) according to the manufacturer’s instructions. TE1 cells were lysed in RIP buffer consisting of 150 mM KCL, 25 mM Tris-HCl (pH 7.4), 5 mM EDTA, 5 mM DTT, 0.5% Triton X-100, supplemented with RNase inhibitor Ribolock and proteinase inhibitor cocktail. The lysate was mixed with SRSF1 antibody (Santa Cruz Biotechnology, USA) or normal IgG coupled beads and left under rotation at 4 °C. Beads were subsequently washed in lysis buffer and the input RNA was purified, and thus analyzed by RT-PCR and qRT-PCR using the primer of DGCR5. RIP assay was performed as we previously described10.

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