GSM8702908: Normal_fibroblasts_IL-5_stimulated_1; Homo sapiens; RNA-Seq
Following 6 hours of treatment with IL-5, Treated and non-treated cells were collected by trypsinization. Cells were pelleted and RNA was extracted from the cell pellet using the Qiagen RNeasy kit according to the manufacturer's instructions.  Genomic DNA removal from all RNA extractions was ensured by treating the RNA with TURBO DNA-free Kit. From Turbo DNase-treated RNA, 100 ng was used for the library preparation. Whole transcriptome sequencing was performed using targeted RNA-Seq with Ion AmpliSeq whole transcriptome human gene expression kit for 20,000 specific human RNA transcripts. In brief, cDNA was synthesized using a SuperScript VILO cDNA Synthesis kit and amplified using Ion AmpliSeq gene expression core panel primers. The amplified products were subjected to enzymatic shearing to get amplicons of ~200 bp, then ligated with the adapter and the unique barcodes. Next, the constructed library was purified using Agencourt AMPure XP Beads, quantified using an Ion Library TaqMan Quantitation Kit, and further diluted to 100 pM, and pooled equally with 16 individual samples. The diluted library was amplified and enriched in Ion Chef System according to the manufacturer's instructions. The prepared template libraries were sequenced on an Ion S5 XL Semiconductor sequencer using Ion 540 Chip. Targeted RNA-Seq using Ion AmpliSeq whole transcriptome human gene expression kit