|
| Status |
Public on Mar 08, 2017 |
| Title |
miR17_92_tKO3_25.5hr |
| Sample type |
SRA |
| |
|
| Source name |
FoB cells_miR17_92_tKO_stimulated
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6 x 129 mixed background
age: 8-week-old
genotype/variation: tKO
cell type: follicular B cells
treatment: stimulated with LPS/IL-4 for 25.5hr in B cell media
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Purified ribosome footprints were extracted using miRNeasy Kit from Qiagen (cat. no. 217004)
RNA libraries were prepared for sequencing using ribosome profiling libary prep protocol (Ingolia et al., Nature Protocol 2012). Indexed footprint library was generated with 4 or 6 cycles of PCR using index library primers, and avoided broad, slower migrating smear bands.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
Ribosome footprints
FoB25.5_TKOrep3
|
| Data processing |
Illumina Casava1.8.2 software used for basecalling.
The raw sequencing data were separated into individual samples using index primer sequences. Then, the miRNA linker and the first nucleotide of 5'-end are clipped and trimmed using fastx toolkit (http://hannonlab.cshl.edu/fastx_toolkit/). Reads obtained from rRNAs are discarded using Bowtie v1.0.0 using 23 seed flag (Langmead et al, 2009). Illumina iGenome (mm10) were used as a reference. The non-rRNA sequencing reads were further aligned to the reference genome (mm10) using TopHat v2.0.11 with added option, "no-novel-juncs"
Only the perfect-match aligned reads were further extracted from accepted hits (accepted_hits.bam) to reduce potential noise derived from relatively short ribosome footprint length (Ingolia et al, 2012). Differential ribosome footprint levels in different genotypes were further quantified using Cuffdiff (v 2.2.1)
To accept only translated genes, we took cut-off of ribosome footprints at fragments per kilobase per million (FPKM) > 1, and only selected genes that were quantified in all three biological replicates.
Genome_build: mm10
Supplementary_files_format_and_content: [.xlsx] tab-delimited file includes FPKM values for each Sample
|
| |
|
| Submission date |
Jun 23, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Changchun Xiao |
| E-mail(s) |
cxiao@scripps.edu
|
| Phone |
(858) 784-7640
|
| Organization name |
The Scripps Research Institute
|
| Department |
Immunology & Microbial Science
|
| Lab |
Changchun Xiao Lab
|
| Street address |
10550 North Torrey Pines Road
|
| City |
La Jolla |
| State/province |
California |
| ZIP/Postal code |
92037 |
| Country |
USA |
| |
|
| Platform ID |
GPL19057 |
| Series (1) |
| GSE83684 |
Ribosome Profiling analysis of Follicular B (FoB) cells purified from WT, miR-17~92 transgenic and miR-17-92 tKO mice |
|
| Relations |
| BioSample |
SAMN05291452 |
| SRA |
SRX1873953 |
