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Long-read Nanopore sequencing of mouse neural stem cells

ABSTRACT: Purpose: The aim of this study is to identify genes that are under the transcriptional control of the epigenetic modifier Smchd1 in mouse neural stem cells. We profiled the transcriptomes of mouse neural stem cells from samples that were either wild-type or contained a null mutation in the epigenetic regulator Smchd1 using Oxford Nanopore Technologies (ONT) direct cDNA sequencing protocol and a PromethION sequencer.

ORGANISM(S): Mus musculus

PROVIDER: GSE151841 | GEO | 2020/06/18

REPOSITORIES: GEO

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Project description:Purpose: Nanopore sequencing is a new generation of Nanopore-based single-molecule real-time electrical signal sequencing technology for full-length transcriptome sequencing. We use Oxford Nanopore Technology (ONT) three-generation full-length transcriptome sequencing to detect all genetic structural changes in the transcriptome of tumor endothelial cells (TECs) 2H-11 and normal endothelial cells (NECs) SVEC4-10. Methods: The mRNA profiles of three samples of mouse tumor endothelial cells (TECs) 2H-11 and three samples of mouse normal endothelial cells (NECs) were generated by Oxford Nanopore Technologies long read processing, using PromethION. Differential expression analysis of six samples was performed using the DESeq2 R package (1.6.3). Genes with a FDR < 0.01 and foldchange ≥ 2 found by DESeq2 were assigned as differentially expressed. Gene Ontology (GO) enrichment analysis of the differentially expressed genes (DEGs) was implemented by the GOseq R packages. KOBAS software was used to test the statistical enrichment of (DEGs) in KEGG pathways. Results: In mouse tumor endothelial cells 2H-11,1847 genes are up-regulated and 1202 genes are down-regulated. According to the Gene ontology (GO) enrichment analysis of differentially expressed genes (DEGs), we found that different functional trends related to metabolic processes, developmental processes, localization, immune system processes, and locomotion are the main reasons for the differences. DEGs are mainly enriched in signal pathways related to cancer, immunity and metabolism, involving Pathways in cancer, Antigen processing and presentation, Proteoglycans in cancer, Focal adhesion, MAPK signaling pathway, Protein digestion and absorption, ECM-receptor interaction, PI3K-Akt signaling pathway and Glutathione metabolism. We also obtained the structural variation of transcripts such as alternative splicing, gene fusion, and alternative polyadenylation and accurately quantified the expression of the transcript. Conclusions: Differentially expressed genes (DEGs) in key pathways may be potential diagnostic markers or therapeutic targets of TECs. Our study could help better understand the molecular mechanisms of tumor endothelial cells (TECs) involved in tumorigenesis and development. And our data also provide useful genetic resources for improving the genome and transcriptome annotations of TECs and normal endothelial cells (NECs).

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Long-read Nanopore sequencing of mouse neural stem cells

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