title: GSM8262425: MainExperiment_GE_A549cells_4weeks_control_rep1; Homo sapiens; RNA-Seq
library_construction_protocol: RNA was isolated from the cells using the RNeasy Mini Kit (Qiagen, Hilden, Germany); The concentration and integrity of the isolated RNA was quantified using the NanoDrop (Thermo Fisher Scientific) and Agilent TapeStation. A total of 500 ng RNA for each sample with RNA integrity number (RIN) values > 8 was used for sequencing library construction. PolyA RNA was first purified via NEBNext Poly(A) mRNA Magnetic Isolation Module (New England BioLabs). The polyA RNA was employed as the input for the preparation of the library for RNA-seq, which was conducted using the NEBNext Ultra II Directional RNA Library Prep kit (New England BioLabs). The approach uses dUTP-based method to preserve strand information. After indexing via PCR (10 cycles) enrichment, resulting transcriptome libraries were quantified by Qubit DNA Assay (Thermo Fisher Scientific) and their fragment size distribution was estimated by TapeStation D1000 ScreenTape (Agilent). The libraries were then pooled/multiplexed at an equimolar amount each and loaded onto Illumina NovaSeq 6000 paired-end sequencing (2 × 150 bp), yielding approximately 20M PE reads per sample. Library preparation and sequencing were conducted by Azenta Life Sciences (Tokyo, Japan).