RNA-seq of of doxorubicin exposed SV40 immortalized human cardiomyocytes with FOXO3 knock-down or shGFP control
RNA-seq of of doxorubicin exposed SV40 immortalized human cardiomyocytes with FOXO3 knock-down or shGFP control
After the exposure, cells were washed once with PBS and subsequently lysed in 0.5 mL QIAzol Lysis Reagent (79306, Qiagen). Total RNA was extracted using chloroform and isopropanol precipitation. Poly(A) libraries were generated using the Zephyr G3® NGS automated system with the NEXTFLEX® Rapid Directional RNA-Seq Kit 2.0 (NOVA-5198–02, PerkinElmer). Library preparation included NEXTFLEX® Poly(A) Beads 2.0 (NOVA-512992, PerkinElmer) for mRNA enrichment and NEXTFLEX® Unique Dual Index Barcodes (NOVA-512923, PerkinElmer) for sample multiplexing. Each library was prepared using 100 ng of total RNA as input, with RNA integrity numbers (RIN ≥ 8). A total of 13 PCR cycles were performed during library amplification. 