GSM8546112: outer hair cells, tamoxifen p2-5, rep 2; Mus musculus; RNA-Seq
The mRNA of Stat3ΔOHC and control  of the organ of Corti was carefully separated from the cochlea and digested with a lysis buffer containing 1mg/mL Collagenase IV , 2mg/mL Papain and 1mg/mL Thermolysin for 10min at 37°C. Subsequently, OHCs were incubated with Slc26a5-antibodyand then with lgG anti-rabbit microbeads for indirect selection of OHCs. The cell suspension was passed through MACS® Separator columns  to enrich magnetically labeled OHCs. After elution of OHCs, the RNA was extracted using the Arcturus Pico Pure Isolation Kit. The RNA samples were checked using 2100 Bioanalyzer with RNA 6000 Pico kit and DNase treatment was performed. cDNA libraries suitable for sequencing were prepared with SMARTer® Stranded Total RNA-Seq Kit v3 - Pico Input Mammalian according to manufacturer's instructions. The PCR amplification was performed using 16 PCR cycles. Libraries were quantified by QubitTM dsDNA HS Assay Kit and quality was checked using 2100 Bioanalyzer with High Sensitivity DNA kit before equimolar pooling. Sequencing of pooled libraries was performed in paired-end mode with 75 nt read length on the NextSeq 500 platform (Illumina).