GSM2176507: total input RNA, ribo-zero rep2; Homo sapiens; RIP-Seq
The total cellular RNA was extracted by TRI reagent according to manufacturer's instruction. For the extraction of total SIRT7-RIP RNA, we performed RIP (RNA immunoprecipitation) experiment. Briefly, the lysates were incubated with anti-Flag M2 affinity gel to immunoprecipitate SIRT7 ribonucleoprotein complex (RNP) . Anti-Flag beads were washed extensively and then subjected to proteinase K digestion to release RNA from SIRT7 RNP. RNA associated with SIRT7 was then extracted by standard phenol-chloroform extraction followed with ethanol precipitation. Ribosomal RNA was substracted from both input cellular RNA and SIRT7-RIP RNA during library construction with the Ribozero Gold HMR kit (Illumina). Directional RNA-seq libraries were prepared from 25-100ng rRNA-subtracted RNA using the NEBNext Ultra Directional RNA Library Prep Kit for Illumina (New England Biolabs).
GEO Accession
GSM2176507