GSM9149235: CTRLin26JUN; Homo sapiens; RNA-Seq
Following polysomal profiling, RNA from total/input and polysome associated fractions was isolated using Qiagen AllPrep RNA/Protein Kit, Cat#: 80404 according to manufacturers protocol. RNA samples obtained from polysome profiling experiments were sent to Novogene Corporation for processing. RNA quantity, integrity, and purity were assessed using the Agilent 5400 Fragment Analyzer System. Samples with an RNA Integrity Number (RIN) above 6.0 qualified for library preparation. Messenger RNA (mRNA) was purified from total RNA using poly-T oligo-attached magnetic beads. Following fragmentation, first-strand cDNA synthesis was performed using random hexamer primers, followed by second-strand cDNA synthesis. Library preparation included end repair, A-tailing, adapter ligation, size selection, amplification, and purification. The final libraries were assessed using a Qubit fluorometer and real-time PCR for quantification, as well as a Bioanalyzer for size distribution analysis. Quantified libraries were pooled and sequenced on Illumina platforms based on the effective library concentration and desired data output. Quantified libraries were pooled and sequenced on the Illumina NovaSeq X Plus (25B flowcell), using a paired-end 150bp (PE150) sequencing strategy. The clustering of the index coded samples was performed according to the manufacturer's instructions.