|
| Status |
Public on Jul 15, 2024 |
| Title |
RNA-Seq Beta-cells_Y_3 |
| Sample type |
SRA |
| |
|
| Source name |
Pancreas
|
| Organism |
Mus musculus |
| Characteristics |
tissue: Pancreas
cell type: Beta cells
genotype: C57/Bl6 background
treatment: 2 months
|
| Growth protocol |
The tissues of 5 healthy animals were pooled per-replicate for cell isolation, with 3-5 replicates (totalling 15-25 animals) collected per cell type/condition. Multi-omic bulk profiling (OMNI-ATAC-seq assay and RNA-seq) was performed on purified populations collected via fluorescence activated cell sorting (FACS).
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
RNA-seq
Poly-A based RNA sequencing libraries were generated as described by us (Sun et al. Nat Comms 2021; https://doi.org/10.1038/s41467-021-22863-0.). In brief, a 3-prime biased approach was used to generate sequencing libraries. An 8 bp sample index and a 10 bp unique molecular identifier (UMI) were added during initial poly(A) priming and pooled samples were amplified using a template-switching oligonucleotide. The Illumina P5 (5′-AAT GAT ACG GCG ACC ACC GA-3′) and P7 (5′-CAA GCA GAA GAC GGC ATA CGA GAT-3′) sequences were added by PCR and Nextera transposase, respectively. The library was designed so that the forward read (R1) utilises a custom primer (5′-GCC TGT CCG CGG AAG CAG TGG TAT CAA CGC AGA GTA C-3′) to sequence directly into the index and then the 10 bp UMI. The reverse read (R2) uses the standard R2 primer to sequence the cDNA in the sense direction for transcript identification.
For RNA sequencing, a 3-prime biased approach was used to generate sequencing libraries as described previously (Sun et al. Nat Comms 2021; https://doi.org/10.1038/s41467-021-22863-0.). Barcoded 19bp R1 and 111bp R2 paired-end reads per sequencing lane were assigned to individual samples using the sabre software (version 80faf94) with options “-u –m2 –l 10’ (https://github.com/najoshi/sabre) (Tsyganov et al. JOSS 2018; https://doi.org/10.21105/joss.00583).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
DNBSEQ-G400 |
| |
|
| Description |
Beta-cells_Y_3
|
| Data processing |
Demultiplexed single end reads per sample here presented were mapped to the mouse GRCm38.p6/mm10 genome primary assembly using Spliced Transcripts Alignment to a Reference (STAR) software version 2.5.2b (Dobin et al. Bioinformatics 2013; https://doi.org/10.1093/bioinformatics/bts635). STAR was run with mode alignReads with parameters “--outFileterMatchMinOverLred 0.3 --outFilerScoreMinOverLread 0.3 --twopassMode Basic –sjdbGTFfile –quanMode GeneCounts”. Reads with duplicated UMIs were removed using the software Je version 2.0.RC option markdupes (Girardot et al. BMC Bioinformatics, 2016; https://doi.org/10.1186/s12859-016-1284-2). Transcript quantification was performed using featureCounts version 2.0.1 with parameters “–Q10 -M" (exonic regions of GENCODE’s vM24 annotation version) (Liao et al. NAR 2019; https://doi.org/10.1093/nar/gkz114).
Assembly: mm10
Supplementary files format and content: (.txt) counts
|
| |
|
| Submission date |
Jul 03, 2024 |
| Last update date |
Jul 15, 2024 |
| Contact name |
Christian Nefzger |
| Organization name |
The University of Queensland
|
| Department |
Institute for Molecular Biology
|
| Lab |
Cellular Reprogramming and Ageing
|
| Street address |
306 Carmody Road
|
| City |
Brisbane |
| State/province |
Queensland |
| ZIP/Postal code |
4067 |
| Country |
Australia |
| |
|
| Platform ID |
GPL28457 |
| Series (2) |
| GSE223049 |
Bulk RNA-seq in young (2 months) and aged (22-24 months) mice across 23 cell-types |
| GSE223050 |
The activity of early-life gene regulatory elements is hijacked in aging through pervasive AP-1–linked chromatin opening |
|
| Relations |
| BioSample |
SAMN42300467 |
| SRA |
SRX25203028 |
