This record contains raw data related to the article “Evaluation of Oxford Nanopore MinION RNA-seq performance for human primary cells". Abstract: Transcript sequencing is a crucial tool for gaining a deep understanding of biological processes in diagnostic and clinical medicine. Given their potential to study novel complex eukaryotic transcriptomes, long-read sequencing technologies are able to overcome some limitations of short-read RNA-seq approaches. Oxford Nanopore Technologies (ONT) offers the ability to generate long-read sequencing data in real-time via portable protein nanopore USB devices at a reasonable cost. This work aimed to provide the user with the number of reads that should be sequenced, through ONT MinION platform, to reach the desired accuracy level for a human cell RNA study. We sequenced three cDNA libraries prepared from poly-adenosine RNA of human primary fibroblasts. Since the runs were comparable, they were combined in a total dataset of 48 million reads. Synthetic datasets with different sizes were generated starting from the total and analyzed in terms of the number of identified genes and their expression levels. As expected, an improved sensitivity was obtained increasing the sequencing depth, in particular for the non-coding genes. The reliability of expression levels was assayed by i) comparison with PCR quantifications of selected genes and ii) by the implementation of a user-friendly multiplexing method in a single run.

This work was supported by the Italian Ministry of Health funds (Ricerca Finalizzata: GR-2018-12366423; ERA-CVD: PICASSO-JTC-2018-042) and by Fondazione Gigi e Pupa Ferrari ONLUS (FPF-14).