GSM5709341: REST_KO_SC_1; Mus musculus; RNA-Seq
Mice were weighed and then sacrificed by cervical dislocation. Relevant tissues were snap frozen in liquid nitrogen and stored at -80°C. Prior to protein or RNA extraction, tissues were ground to a powder using a pestle and mortar pre-chilled with liquid nitrogen. For protein extraction, tissue powders were homogenized using TissueRuptor II (QIAGEN) in RIPA buffer. The homogenates were centrifuged at 13,000xg for 10 minutes at 4°C and the supernatant was collected and quantified with the Bradford colorimetric reaction (Biorad). Total RNA was isolated from tissue powder using QIAzol Lysis Reagent (Qiagen)/chloroform extraction followed by spin-column purification (RNeasy Mini Kit; Qiagen). 15' minute on-column DNase treatment was performed, according to manufacturer's instructions. Illumina "Truseq Stranded mRNA" kit was used to prepare cDNA libraries for Lhx1os +/+ and Lhx1os -/- mice spinal cord samples