RNA seq iPSC various reprogramming strategy
Total RNA extraction was performed on 6 hiPSC lines and on Tie2+ NPP cells of the 3 heathy donors using the RNeasy kit (#74104, Qiagen) with on-column DNase digestion with the RNAse-Free DNase Set (#79254, Qiagen) following the original producer's instructions. RNA Screen Tape (Agilent 4150 TapeStation system) was used to assess RNA integrity. Bulk mRNA sequencing in paired-end 150 bp was performed on BGISEQ sequencing platform by BGI Genomics. Filtered clean reads were aligned to the genome version reference sequence: homo_sapiens_9606.NCBI.GCF_000001405.40_GRCh38.p14.v2201, by HISAT, and then mapped to the reference genes using Bowtie2. Gene expression quantification was performed using RSEM software (v1.3.1) and heatmaps of gene expression clustering between different samples were generated using pheatmap (v1.0.8). DESeq2 (v1.4.5) was used for differential gene detection.