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Circadian bulk RNA-seq profiling in human cerebral organoids

ABSTRACT: Bulk mRNA sequencing was performed on human cerebral organoids derived from gene-edited H9 human embryonic stem cells to characterise circadian transcriptomic dynamics. Organoids were collected every 3 hours across 17 timepoints over a 48-hour period (GCT0–GCT48) under constant conditions, with three biological replicates per timepoint. Libraries were prepared using poly(A)+ selection with the KAPA mRNA HyperPrep kit (KK8581) and sequenced using single-end 70 bp reads on an Illumina NextSeq 2000 platform, generating approximately 40 million reads per sample. The resulting dataset supports analyses of circadian mRNA expression and underlies the transcriptomic analyses reported in the manuscript ‘Circadian clocks time targets of disease in human brain organoids’.

ORGANISM(S): Homo sapiens

PROVIDER: GSE317981 | GEO | 2026/09/20

REPOSITORIES: GEO

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Project description:Purpose: The goal of this study are to investigate the TRPM1-regulated genes using RNAseq to compare the transcriptome profiling between 661W cells expressing TRPM1 and control vectors Methods: RNAs were isolatedusing the RNeasy Mini kit (Qiagen). RNA-seq libraries were prepared using the KAPA mRNA HyperPrep Kit (KAPA Biosystems, Roche, Basel, Switzerland) and validated using the Qsep 100 DNA/RNA Analyzer (BiOptic Inc., Taiwan). Libraries were sequenced on a NovaSeq 6000 sequencer (Illumina, CA, USA). Clean reads were aligned to the mouse genome (GRCm38) using HISAT2 (version 2.1.0) after removing low-quality reads. The differential expression of genes between TRPM1-overexpression and control cells was computed using the fragments per kilobase of transcript per million mapped reads calculated by featureCounts (version 2.0.0). Raw read counts were imported into edgeR (version 3.28.1) and analyzed by using R package of DESeq (version 1.40.0). Genes with false discovery rate (FDR) p-value < 0.05 adjusted by using Benjamini–Hochberg (BH) method were considered as differentially expressed genes (DEGs). Gene set enrichment analysis of the genes differentially expressed upon TRPM1 expression was done using the Gene Set Knowledgebase（GSKB）hallmark gene sets. Results: We had 40,922,040 clean reads in the control group and 44,244,608 clean reads in the TRPM1-overexpressing group. We mapped 43,253, 668 (97.76%) sequence reads in the control group and 39,942,649 (97.6%) sequence reads in the TRPM1-overexpressing group. We identified 16,014 transcripts. 76 transcripts showed differential expression between the vector control group and TRPM1-expressing group, with a fold change ≥1.5 and p value <0.01. Gene set enrichment analysis of the genes differentially expressed upon TRPM1 expression uncovered several TRPM1-regulated genes that may contribute to photoreceptor function, such as retina morphogenesis and JAK-STAT cascade. Conclusions: Our study represents the genes associated with TRPM1 overexpression in 661W photoreceptor cells using RNA-seq approach. The overexpression of TRPM1 may contribute to regulate the photoreceptor morphogenesis and function

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Circadian bulk RNA-seq profiling in human cerebral organoids

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