GSM7122447: SMCTRAP-Athero_121_Input; Mus musculus; RNA-Seq
Mice were euthanized in CO2 chamber (Metos) and the tissues were dissected while the mice were kept on ice after perfusing with ice-cold 1X PBS (Gibco). Aorta was snap-frozen using liquid nitrogen for TRAP-Seq. The frozen aortic tissue was pulverized using a TissueLyser II bead mill (QIAGEN) and added with low salt lysis buffer for homogenization. 5% of lysate was used for the bulk RNA extraction and the remaining lysate was immunoprecipitated with anti-GFP clone 19F7 and clone 19C8 antibodies. The Biotinylated Protein L (0.5 mg; Pierce #29997) and Dynabeads MyOne Streptavidin T1 (10mg/ml; Invitrogen 65602) are mixed to form bead-protein complex and capture the GFP antibodies in the samples. RNA from the lysate was extracted using RNeasy Micro Plus kit (QIAGEN, 74034). The SMARTer Stranded Total RNA-Seq Kit v2 - Pico Input Mammalian (TaKaRa, 634411, 634412) was used for library preparation, followed by single-end 76-bp sequencing on an Illumina NextSeq 500 instrument.