|
| Status |
Public on Mar 17, 2023 |
| Title |
Y3_240_RNA |
| Sample type |
SRA |
| |
|
| Source name |
mouse liver
|
| Organism |
Mus musculus |
| Characteristics |
tissue: liver
strain: C57BL/6JN
group: young
time point: 240
|
| Treatment protocol |
Old as treated group
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated from frozen tissue by homogenization in Trizol followed by isopropanol precipitation. The RNA was further purified using RNeasy columns (Qiagen). An on-column DNase I digestion was performed during the purification step to remove genomic DNA. The RNA amount and integrity were confirmed using the Qubit RNA HS Assay Kit and RNA IQ Assay (Thermo Fisher) respectively.
~Total RNA (~1 µg) was used to make RNA-seq libraries following the Zymo-Seq Ribo-free Total RNA Library Kit (Zymo Research) instructions with dual indexing. Prior to ribo-depletion, total RNA from young livers were spiked with 2 µl of a 1:100 dilution of ERCC Ex-fold mix 1 while that from old livers were spiked with 2 µl of 1:100 dilution of ERCC Ex-fold mix 2 (Thermo Fisher). Library quality and quantity was confirmed on a BioAnalyzer (Agilent) DNA HS chip. Equimolar amounts of each library were combined, and the pooled library was further quantified using a NEBNext Library Quant Kit (New England Biolabs).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
NextSeq 550 |
| |
|
| Data processing |
Illumina sequencing reads (~32 million paired-end reads per sample) were de-multiplexed using bcl2fastq/2.20.0
Reads were trimmed to remove adapter sequences using trimmomatic/0.39. The quality of the resulting FASTQs was assessed using FASTQC/0.11.9 and reads were aligned to the mouse reference genome (assembly GRCm38/mm10, concatenated with the ERCC transcripts) using STAR/2.7.5b.
BAM files were sorted and indexed using samtools/1.10, and duplicates were removed using picard/2.20.8. The BAM files were then filtered to retain alignments with a minimum mapping quality of 10 using samtools/1.10.
The bamCoverage option in deepTools/3.5.0 was used to generate RPKM normalized bigWig files. H3 and input reads were subtracted from H3K27me3 and IgG respectively with bigwigCompare.
The featureCounts function of the Rsubread R package/2.6.4 was used to estimate counts for all genes and ERCC transcripts.
Genome_build: mm10
Supplementary_files_format_and_content: bigwig files were generated using bamCoverage in deepTools/3.5/0; Scores represent the coverage on the genome location
Supplementary_files_format_and_content: count file was generated with featureCounts function of the Rsubread R package/2.6.4.
|
| |
|
| Submission date |
Oct 12, 2021 |
| Last update date |
Mar 19, 2023 |
| Contact name |
Supriyo De |
| Organization name |
NIA-IRP, NIH
|
| Department |
Laboratory of Genetics and Genomics
|
| Lab |
Computational Biology & Genomics Core
|
| Street address |
251 Bayview Blvd
|
| City |
Baltimore |
| State/province |
Maryland |
| ZIP/Postal code |
21224 |
| Country |
USA |
| |
|
| Platform ID |
GPL21626 |
| Series (2) |
| GSE185705 |
A hyper-quiescent chromatin state is a barrier to productive regeneration during aging. [RNA-Seq] |
| GSE185708 |
A hyper-quiescent chromatin state is a barrier to productive regeneration during aging. |
|
| Relations |
| BioSample |
SAMN22224067 |
| SRA |
SRX12575586 |
