empty vector transfected SH-SY5Y 1
For each sample, 1 g RNA was used for sequencing library construction with the NEBNext UltraTM RNA Library Prep Kit for Illumina (NEB, USA) according to the manufacturers instructions. Index codes were added to attribute sequences to each sample. Index-coded samples were clustered by TruSeq PE Cluster Kit v3-cBot-HS (Illumia) on a cBot Cluster Generation System and sequenced on an Illumina Novaseq platform and 150 bp paired-end reads were obtained. Paired-end clean reads were then aligned to the reference genome using Hisat2 v2.0.5. The mapped reads of each sample were assembled by StringTie (v1.3.3b) using a reference-based approach. FeatureCounts v1.5.0-p3 was used to calculate the fragments per kilobase of transcript sequence per million base pairs sequenced (FPKM) of each gene. The read counts were adjusted using the edgeR R package (3.18.1) through one scaling normalized factor for each sequenced library. Differential expression analysis of two conditions was performed using the edgeR R package. P values were adjusted using the Benjamini and Hochberg method. A corrected P-value < 0.05 and absolute fold change of 2 were set as the threshold for significantly differential expression. Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), DO (Disease Ontology), and DisGeNET enrichment analysis of differentially expressed genes were performed using the clusterProfiler R package.