RNAseq analysis of ClpP siRNA treated mouse primary hepatocytes

We report the gene expression profile in mouse primary hepatocytes after treatment of ClpP siRNA. Overall design: To analyze the changes of gene expression after treatment with ClpP siRNA in mouse primary hepatocytes, the mouse primary hepatocytes were treated with ClpP siRNA using Lipofectamine 2000. The TruSeq RNA Sample Preparation Kit v2 (Illumina, #RS-122-2001) was used to convert poly-A containing mRNAs in total RNA to a cDNA library using poly-T oligos linked to magnetic beads. Following mRNA purification, the RNA was physically fragmented at an elevated temperature prior to reverse transcription and cDNA generation. The fragmentation step resulted in the production of an RNA-Seq library containing inserts ranging in size from approximately 100-400 base pairs (bp). The average insert size in an Illumina TruSeq RNA-Seq library is approximately 200 bp. The cDNA fragments were then subjected to an end-repair process, during which a single 'A' base was added to the 3' ends and then adaptors were ligated. Finally, the products were purified and enriched by PCR to generate the final double-stranded cDNA library. The libraries were quantified using KAPA Library Quantification Kits for Illumina Sequencing platforms according to the qPCR Quantification Protocol Guide (KAPA Biosystems, KK4855) and qualified using the TapeStation D1000 ScreenTape device (Agilent Technologies, 5067-5582).