 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Oct 21, 2024 |
| Title |
wild type_BiolRep2 |
| Sample type |
SRA |
| |
|
| Source name |
cell culture
|
| Organism |
Schizosaccharomyces pombe |
| Characteristics |
cell type: cell culture
genotype: 972h- wild type
treatment: pull down with Dynabeads (Invitrogen, magnetic Streptavidin) and eluted with biotin elution buffer (12.5 mM biotin – Invitrogen, 7.5 mM HEPES pH 7.5, 75 mM NaCl, 1.5 mM EDTA, 0.15% SDS, 0.075% sarkosyl, 0.02% Na-Doc)
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted with QIAzol and RNeasy Mini kit (Qiagen) according to the manufacturer’s recommendations, including a DNase treatment.
RNA quality was assessed in an Agilent 2100 Bioanalyzer and strand-specific cDNA libraries were made NEXTflex Rapid Directional qRNA-Seq Kit (PerkinElmer, formerly BiooScientific). Libraries were quantified in a Qubit, pooled and 4 nM of the pooled library was sequenced
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina MiSeq |
| |
|
| Data processing |
Libraries were spiked with 20% PhiX (Illumina) to increase the complexity of the libraries to accommodate clustering and sequenced in an Illumina MiSeq (75 bp, paired-end) instrument.
All quality control steps including quality filtering, demultiplexing and adapter removal were performed with Illumina BaseSpace in-house tools. The quality of the sequences was confirmed with FastQC (ver 0.11.2, https://www.bioinformatics.babraham.ac.uk/projects/fastqc/).
Reads were mapped with Hisat2 (version 2.2.1, code used: hisat2 -x genomeIndex --rna-strandness FR --trim5 8 -1 pair1.fastq.gz -2 pair2.fastq.gz -S out.sam --summary-file summary.txt)
sam files were converted to bam files and sorted/indexed with samtools (ver 1.14) and sorted bam files were used to count reads per gene with htseq (ver. 0.12.3, https://htseq.readthedocs.io/en/master/index.html; code use: htseq-count -s reverse -f bam -t gene -i ID file.bam annotation.gff3 > file.count).
The top aal1-bound RNAs were determined with edgeR (version 3.32.1; Since there were no significantly enriched RNAs (FDR ≤0.05) in the wild-type relative to aal1∆ cells, we chose the top 30 enriched RNAs which were verified in IGV (ver 2.8.0, https://software.broadinstitute.org/software/igv/). For IGV inspection, strand-specific MiSeq reads were processed further with deepTools (ver 3.5.1, https://deeptools.readthedocs.io/en/develop/) to get normalised gene coverage in counts per million (cpm) per 50 bp bins (bam to bigWig files) for direct comparison.
Assembly: genome build as of January 2020 downloaded from pombase.org
Supplementary files format and content: raw gene counts files combined as a table with counts for each biological replicate as a separate column. In .xlsx format (excel sheet)
Supplementary files format and content: bigwig (.bw) files with normalised gene coverage in counts per million (cpm) per 50 bp bins (bam to bigWig)
Library strategy: ChIRP-RNAseq
|
| |
|
| Submission date |
Sep 13, 2023 |
| Last update date |
Oct 21, 2024 |
| Contact name |
Shajahan Anver |
| E-mail(s) |
s.anver@ucl.ac.uk
|
| Organization name |
University College London
|
| Department |
Genetics, Evolution and Environment
|
| Street address |
Gower Street London
|
| City |
London |
| ZIP/Postal code |
WC1E 6BT |
| Country |
United Kingdom |
| |
|
| Platform ID |
GPL16192 |
| Series (2) |
| GSE243035 |
Ageing-associated long non-coding RNA extends lifespan and reduces translation in non-dividing cells [aal1_ChIRP-RNAseq] |
| GSE243036 |
Ageing-associated long non-coding RNA extends lifespan and reduces translation in non-dividing cells |
|
| Relations |
| BioSample |
SAMN37377068 |
| SRA |
SRX21765509 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7777757_ChIRP-RNAseq_wt2_coverage.bw |
178.0 Kb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
|
|
|
|
 |
