GSM7899051: Muscle, air (M2R1_S17); Rattus norvegicus; RNA-Seq
Uniform portions of frozen tissue were sectioned  and processed with RNeasy non-fibrous, fibrous, or lipid tissue mini kits (Qiagen, Valencia, CA) following manufacturer directions (kit dependent upon tissue type). RNA quantity and purity (260/230 and 260/280 ratios) were spectrophotometrically quantified using a Nanodrop 1000 (ThermoFisher Scientific Inc., Waltham, MA). RNA integrity was assessed by the RNA 6000 LabChip® kit and a 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA) using a RIN cutoff of 7. Samples processing for mRNA sequencing was performed on the Apollo324 automated system (Takara Bio Inc., Kusatsu, Japan) for library prep with PrepX mRNA 48 protocol v19, using the PrepXTM RNA-seq for Illumina Library Kit (Takara), SuperScript III reverse transcriptase (ThermoFisher), and AMPure XP Beads (Agilent). PCR amplification with 48 index primers was run for 16 cycles and resulting PCR product quality was analyzed again via Qubit (ThermoFisher) and bioanalyzer. An RNA-seq library was prepared with Wafergen's PrepX mRNA 48 protocol and dsDNA products were prepared from cDNA. Each library was sequenced according to Illumina NextSeq 500, with a final concentration of 2.2 pM + 2% PhiX and run for 75 cycles (Illumina Inc., San Diego, CA).