|
| Status |
Public on Oct 12, 2018 |
| Title |
rat_rnaseq_oligo_2 |
| Sample type |
SRA |
| |
|
| Source name |
nuclear RNA from specific cell types obtained via the INSPECTION method
|
| Organism |
Rattus norvegicus |
| Characteristics |
strain: Sprague Dawley
brain region: cerebellum
cell type: Oligodendrocyte
target molecule: nuclear RNA
|
| Growth protocol |
All procedures involving mice were approved by The Rockefeller University Institutional Animal Care and Use Committee (IACUC) and were in accordance with the National Institutes of Health guidelines. Mice were housed in a specific-pathogen free facility, in groups of four to five per cage under a 12-h light-dark cycle with food and water ad libitum. Animals were sacrificed between 2-4 months of age.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cerebellum was dissected and cell-type specific nuclei were isolated using the INSPECTION method. Isolated nuclei were resuspended in Buffer PKD (INSPECTION) for RNA purification using the QNeasy FFPE kit. Standard protocols were followed except with the following modifications: after Proteinase K digestion, RNA was spun at max speed for 15 minutes. The supernatant was removed and incubated at 65oC for 30 minutes, 70oC for 30 minutes, and 80oC for 15 minutes. An on-column DNase digestion was performed in place of the in-solution DNAase digestion. RNA quality was determined using Agilent 2100 Bioanalyzer.
Purified RNA was converted to cDNA and amplified using the Nugen Ovation RNA-seq System V2. Amplified cDNA was fragmented, end-repared, adpater ligated, fragment enriched, and sequenced on an Illumina NextSeq 500 instrument.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Data was mapped to the Rattus norvegicus reference genome (rn6).
Analysis for RNA-seq datasets were performed as described in Xu et al., 2017
RNA-seq: Reads were aligned to the rn6 genome using STAR v2.4.2a (--outFilterMismatchNmax 999 --outFilterScoreMinOverLread 0 --outFilterMatchNminOverLread 0 --outFilterMatchNmin 35 --outFilterMismatchNoverLmax 0.05)
RNA-seq: Gene expression levels were estimated using featureCounts v1.5.2 (-C) using annotation files for whole genes or exons
RNA-seq: Visualization tracks were generated using igvtools v2.3.32.
Genome_build: rn6
Supplementary_files_format_and_content: Tab delimited txt format; Table of raw counts for all samples generated using species-specific annotations, mouse-rat-human ortholog annotations, or mouse-human ortholog annotations.; tdf format for visualization in IGV.
|
| |
|
| Submission date |
Jul 26, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Xiao Xu |
| E-mail(s) |
[email protected]
|
| Organization name |
The Rockefeller University
|
| Department |
Laboratory of Molecular Biology
|
| Lab |
Nathaniel Heintz
|
| Street address |
1230 York Ave
|
| City |
New York |
| State/province |
NY |
| ZIP/Postal code |
10065 |
| Country |
USA |
| |
|
| Platform ID |
GPL20084 |
| Series (2) |
| GSE101917 |
Species and Cell-Type Properties of Classically Defined Human and Rodent Neurons and Glia [RAT RNA-Seq] |
| GSE101918 |
Species and Cell-Type Properties of Classically Defined Human and Rodent Neurons and Glia |
|
| Relations |
| BioSample |
SAMN07417151 |
| SRA |
SRX3038930 |
