RNA-seq of primary mouse podocytes with KDM6A overexpression against controls

We performed RNA-seq experiments (three replicates) on primary mouse podocytes that were infected with control lentivirus or KDM6A-encoding lentiviruses. RNAs were first extracted using Trizol reagent (Invitrogen, USA) according to the manufacturer's instruction. The purified RNAs were quantified at OD260nm using a ND-1000 spectrophotometer (Nanodrop Technology, USA) and qualified using a Bioanalyzer 2100 (Agilent Technology, USA) with RNA 6000 LabChip kit (Agilent Technology, USA). All procedures for library preparation and sequencing were performed according to the Illumina protocol. Library construction was carried out using the TruSeq RNA Library Preparation Kit for 75 bp Single-End sequencing on Solexa platform. The sequence was directly determined by sequencing-by-synthesis technology via the TruSeq SBS kit (Illumina Inc., USA). Raw sequences were obtained from the Illumina Pipeline software bcl2fastq v2.0, which was expected to generate 30 million reads per sample. Qualified reads after filtering low-quality data were analyzed using TopHat/Cufflink. Quantification for gene expression was calculated as fragments per kilobase of transcript per million mapped reads (FPKM).