RNA binding of CBFB and hnRNPK

RNA immunoprecipitation assay followed by next generation sequencing (RIPseq) to identify RNAs bound by CBFB and hnRNPK in breast cells. Overall design: One repeat for RIPseq of CBFB and two repeats for hnRNPK. A stable cell line derived from MCF10A cells (ATCC, Cat: CRL-10317) expressing FLAG tagged CBFB was used for CBFB RIP using the FLAG antibody (Sigma, Cat: A2220). Parental MCF10A cells were used for hnRNPK RIP using hnRNPK antibody (Bethyl Laboratory, Cat: A300-674A).

本研究采用RNA免疫共沉淀结合高通量测序（RNA immunoprecipitation assay followed by next generation sequencing，RIPseq）技术，鉴定乳腺细胞中CBFB与hnRNPK所结合的RNA。总体实验设计：CBFB的RIPseq设置1次生物学重复，hnRNPK的RIPseq设置2次生物学重复。CBFB的RIP实验采用源自MCF10A细胞（ATCC，货号：CRL-10317）的稳定表达FLAG标签融合CBFB的细胞系，使用FLAG抗体（Sigma，货号：A2220）进行免疫沉淀；hnRNPK的RIP实验则采用亲本MCF10A细胞，使用hnRNPK抗体（Bethyl Laboratory，货号：A300-674A）完成免疫沉淀。