GSM8772957: A15_9_4_3; Homo sapiens; RNA-Seq
Short term gene changes were assessed in MSCs seeded on tissue chips on 4, 234 and 234F ECM combinations at 150kPa and 900 kPa, following the proliferation protocol. After 48 hours in growth media, cells were lysed and RNA was extracted using GeneJET RNA isolation kit (Fisher Scientific) followed by RNA sequencing by Novogene Corporation (n=4). CASAVA base recognition (Base Calling) was used to convert original image data files from Illumina sequencing platform to sequenced reads in FASTQ format. A series filtration criteria was applied based on uncertain nucleotides constituting more than 10% of either read (n>10%) and with low-quality nucleotides (base quality less than 5) constituting more than 50 per cent of the read. Next, alignment of raw reads was performed to reference genome using HISAT2. To estimate gene expression levels, the fragments per kilobase of transcript sequence per million base pairs sequenced (FPKM) was quantified for each aligned read. This accounted for both sequencing depth and gene length on counting of fragments.