title: GSM9232539: Hep-G2 cell, HiFi FaDe Cas9 editing treatment, 24 hours, replicate 3; Homo sapiens; RNA-Seq
library_construction_protocol: Total RNA was isolated from 5 × 10⁶ cells using a lysis buffer mix containing Proteinase K from the RNAdvance Tissue kit (Beckman Coulter). Cells were vortexed thoroughly and incubated at 37°C for 25 minutes. RNA isolation, including a 15-minute DNase I treatment, was carried out according to the manufacturer's protocol on a Biomek i7 Hybrid robotic workstation (Beckman Coulter). RNA quantity and quality were assessed using a Fragment Analyzer 5300 system (Agilent), and RNA integrity number (RIN) values were recorded. Total RNA-seq libraries were prepared using the KAPA RNA HyperPrep with RiboErase kit (Roche) on a Tecan Fluent® liquid handler, following the manufacturer's instructions. Library integrity and quality were evaluated with the SS NGS Fragment Kit (1–6000 bp; Agilent) on a fragment analyzer. Sequencing was performed as paired-end 2 × 100 bp on an Illumina NovaSeq 6000 platform using a v1.5 SP Reagent kit (Illumina).