title: GSM8881292: DEL_21D Replicate 1 RNA-Seq; Rattus norvegicus; RNA-Seq
library_construction_protocol: RNA was isolated using TRIzol Reagent (Life Technologies) following the manufacturer's protocol. Briefly, 500 μl of TRIzol reagent were added directly to frozen nerve samples, which were mechanically dissociated using a pestle. The resulting RNA pellet was resuspended in 12 μl of sterile RNase-free water. RNA samples were quantified, and their quality and purity were assessed using the Agilent 5400 system, yielding an RNA integrity number (RIN) greater than 6. For non-directional library construction, mRNA was purified from 400 ng of total RNA using poly-T oligo-attached magnetic beads. First-strand cDNA synthesis was performed using random hexamer primers, while the second strand was synthesized using dTTP. Post-reverse transcription steps included end repair, A-tailing, adapter ligation, size selection, amplification, and purification. The library was checked using Qubit and Real-Time PCR for quantification and a bioanalyzer for size distribution detection. Quantified libraries were pooled and sequenced on Illumina platforms based on effective library concentration and data amount.