GSM6327078: Treg, Il10 expressing, rep2 [RNA_G2_pos]; Mus musculus; RNA-Seq
Cells sorted directly into Trizol LS; phase separation was induced with chloroform. RNA was precipitated with isopropanol and linear acrylamide and washed with 75% ethanol. The samples were resuspended in 13 µL nuclease-free water. After RiboGreen quantification and quality control by Agilent BioAnalyzer, 122-400pg total RNA with RNA integrity numbers ranging from undetectable to 9 underwent amplification using the SMART-Seq v4 Ultra Low Input RNA Kit (Clonetech catalog # 63488), with 12 cycles of amplification. Subsequently, 4ng of amplified cDNA was used to prepare libraries with the KAPA Hyper Prep Kit (Kapa Biosystems KK8504) using 8 cycles of PCR. Samples were barcoded and run on a HiSeq 4000 in a PE50 run using the HiSeq 3000/4000 SBS Kit (Illumina).