Illumina HiSeq 2500 paired end sequencing; Cuproptosis-related genes signature and validation of differential expression and the potential targeting drugs in temporal lobe epilepsy
Cuproptosis-related genes signature and validation of differential expression and the potential targeting drugs in temporal lobe epilepsy
Epileptic focus and control tissues were collected from the Department of Neurosurgery. Informed consent was obtained from all patients. Finally, six epileptic focus specimens were obtained from the hippocampus of patients diagnosed with intractable TLE according to the International League Against Epilepsy (ILAE) diagnostic criteria (Blümcke et al., 2013). Because the hippocampal specimens from age matched controls were sparse, we only collected two control hippocampi from autopsied individuals without a known history of neurologic or psychiatric disease, ensuring a 3:1 disease-to control match.No statistical difference was observed in age or sex between TLE patients and controls. All collected samples were immediately placed in centrifugetubes containing TRIzol and stored at −20 °C until total RNAextraction. Total RNA was isolated from the sample homogenate,according to the manufacturer's instructions (Invitrogen,United States). Next, the quantity and quality of total RNA wereassessed using a Nanodrop spectrophotometer (NanoDropTechnologies, United States). Ribo-Zero rRNA Removal Kit (Illumina,United States), and RNA was broken into fragments of approximately 200 bp. The fragmented RNAs were then used tosynthesize first- and second-strand cDNA, and adaptor ligation andenrichment with a low-cycle were performed as instructed by theTruSeq® RNA LT/HT Sample Prep Kit (Illumina, United States).Qubit®2.0 (Life Technologies, United States) and Agilent2,200 TapeStation were used to evaluate the purified library products. Next, they were diluted to 10 pM for in situ cluster generation on a HiSeq2500 pair-end flow cell.