GSM8881289: DEL_14D Replicate 2 RNA-Seq; Rattus norvegicus; RNA-Seq
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.