RNA Seq data of iPSC-derived astrocytes from Progressive Supranuclerar (PSP) patients and controls

Published: 21 April 2023| Version 1 | DOI: 10.17632/wwwrzhbz59.1


RNA seq data of iPSC-derived astrocytes from Progressive Supranuclear Palsy (PSP) patients and controls: number of reads and differentially expressed genes tables. Progressive Supranuclear Palsy patients (PK01, PK02, PK04, PK08, PK09) controls (PK03, PK05, PK06, CQ16)


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Astrocytes were cultured in an astrocyte differentiation medium (AGM, Lonza #3186) for approximately 5 weeks until passage 5. Astrocyte medium was switched to DMEM/F12 without phenol red (Gibco # 21041-025) without fetal bovine serum (FBS) for 48h. Pellets were collected and frozen at -80C until RNA extraction. The total RNA was isolated employing the PureLink RNA mini kit (Thermo Fisher Scientific # 12183020) with DNAse digestion (Thermo Fisher Scientific # 12185010) into the columns. The quality control of total RNA was measured by Bioanalyzer 2100 (Agilent # 5067-1511) and all samples had a RIN between 8 and 9. The isolation of all mRNA content, library preparations, and all NGS conditions were performed according to Illumina´s instructions with TruSeq Stranded mRNA Prep kit. Paired-end sequencing was performed in two runs using the For sequencing, we used NextSeq 500/550 High Output Kit v2.5 (150 Cycles paired-end) kit and NextSeq 550 Instrument from Illumina, Inc. (USA), resulting in an average number of 104.8 million reads. The coverage in the RNA sequencing was around 104.8 million reads per sample, with a Q score >= 30 over 90.%. All sequences in RNA-Seq were aligned to the human genome (GRCh37.75) with the algorithm Spliced Transcripts Alignment to a Reference (STAR v2.7.5c) . The aligned sequences were counted in the RNA-Seq package by the Expectation Maximization package (RSEM v1.3.1) . Data were normalized in the Trimmed Mean of M-values package (TMM) and differential expression was performed in the edgeR package, employing two criteria for up or down gene lists: p-value <0.05 and logFC ≥ 2 or logFC ≤ -2. After sequencing of all transcripts, reads from all samples were previously processed with Trimmomatic tool (v 0.38). Ribosomal sequences were filtered using SortMeRNA (v 2.1) . All sequences were aligned to the human genome GRCh37/hg19 as reference using STAR and the counting of each transcript was performed using RSEM (v1.3.1) The Differentially Expressed Genes (DEGs) were identified when we compared the results of the PSP vs. the control group. The gene expression levels were normalized and calculate using edgeR. Complex Heatmap in the Bioconductor software package for Rstudio (https://www.r-project.org) was used to plot these heatmaps and the distance matrix was used for clustering with Pearson's correlation metric. The threshold for DEGs was established with Log2 Fold change (FC) > 2.0 and p-value <0.05, FDR (False Discovery Rate) <0.001. Samples: Progressive Supranuclear Palsy patients (PK01, PK02, PK04, PK08, PK09) controls (PK03, PK05, PK06, CQ16)


Universidade Estadual de Campinas, Hospital Israelita Albert Einstein, Grupo Hermes Pardini


Neurodegenerative Disorder, RNA, Induced Pluripotent Stem Cell, Astrocyte, Progressive Supranuclear Palsy, Transcriptomics


Fundação de Amparo à Pesquisa do Estado de São Paulo