Fig. 9
From: Advancing brain barriers RNA sequencing: guidelines from experimental design to publication
Overview of the main steps for RNA-Seq data analysis. Raw data goes through quality control steps and, if necessary, pre-processing steps are implemented. Next step is the alignment, most commonly in brain barriers studies being through mapping to reference. One more round of quality control is recommended based on the metrics of the alignment. After sorting, the files can be indexed and visualized in a genome browser. Counting can be attributed at different levels (gene, exon or transcript) and there are multiple algorithms for normalization, both in cases of inter-sample and intra-sample normalization. Finally, after the differential expression analysis, further information can be obtained with steps like gene ontology analysis, gene set enrichment analysis or pathway analysis