GSM9364869: 990705_IL13_5; Mus musculus; RNA-Seq
Tissues/Cells were homogenized in TRIzol and chloroform was used for phase separation. The aqueous phase, containing total RNA, was purified using MagMAX™-96 for Microarrays Total RNA Isolation Kit (Ambion by Life Technologies) according to manufacturer's specifications. Genomic DNA was removed using RNase-Free DNase Set (Qiagen) on the Kingfisher Flex machine (Thermofisher). Experiment 1-2: The strand-specific RNA-seq libraries were prepared from 500 ng RNA using KAPA stranded mRNA-Seq Kit (KAPA Biosystems). The libraries were amplified by twelve-cycle PCR. Sequencing was performed on Illumina HiSeq®2500 (Illumina) by multiplexed single-read run with 33 cycles.  Experiment 3: mRNA was enriched from 4 ug total RNA using two rounds of Dynabeads® Oligo (dT)25 mRNA purification (Invitrogen). Half of the resulting mRNA was converted to mRNA-seq libraries using the ScriptSeq™ mRNA-Seq Library Preparation Kit (EPICENTRE Biotechnologies). The libraries were amplified by twelve-cycle PCR. Sequencing was performed on Illumina HiSeq®2500 (Illumina) by multiplexed single-read run with 33 cycles.