title: RNA sequencing of the DPN treated group
design: This project utilized Oligo(dT) magnetic beads to enrich polyA-structured mRNAs from total RNA, followed by ionic disruption to fragmentize RNA into ~300bp segments. The 300bp length was chosen to balance adapter sequence and valid data ratios, avoiding sequencing hindrances. RNA was then used as a template to synthesize first-strand cDNA with a 6-base random primer and reverse transcriptase, followed by second-strand synthesis. After library construction and PCR amplification, libraries were selected based on size (~450bp), quality-checked with Agilent 2100 Bioanalyzer, and assayed for concentration. Libraries with different Index sequences were mixed proportionally based on effective concentration and data requirements, diluted to 2nM, and transformed into a single-stranded library through alkaline denaturation.