title: GSM8547918: 293Ttotal_SUPV3L1_RIP; Homo sapiens; RIP-Seq
library_name: GSM8547918
library_construction_protocol: The isolated mitochondria, total cells were lysed in TRizol reagent (Life Technologies). RNA from mitochondia, whole cell, mitochondrial fractions and RNA-IP were extracted by using TRizol reagent according to the manufacturer's instructions. For samll RNA seq, the small RNA bands in fractions 2-4 and the input were excised from the gel and RNAs were purified. Small RNA libraries were constructed with NEBNext Multiplex Small RNA Library Prep Set for Illumina following the manufacturer's protocol. For total RNA and mitochondrial RNA sequencing, rRNA is removed from the total RNA sample using TruSeq Stranded Total RNA Library Prep Gold (Illumina), and sequencing libraries were generated using NEB Next Ultra Directional RNA Library Prep Kit for Illumina following the manufacturer's recommendations. The libraries were subjected to 150 nt paired-end sequencing with an Illumina Nova seq 6000 system. Each library generated a depth of ~10 G read pairs. For TRAP1 RIP and ATP5B RIP, sequencing libraries were generated using NEB Next Ultra Directional RNA Library Prep Kit for Illumina following the manufacturer's recommendations. For other RIP-seq, NEB Next Ultra RNA Library Prep Kit was used for library construction of immunoprecipitated RNA. The libraries were subjected to 150 nt paired-end sequencing with an Illumina Nova seq 6000 system. Each library generated a depth of ~10 G read pairs.