|
| Status |
Public on Mar 09, 2016 |
| Title |
U6_sh_hAAT_19nt_rep1_total |
| Sample type |
SRA |
| |
|
| Source name |
liver - total RNA
|
| Organism |
Mus musculus |
| Characteristics |
tissue: liver
strain: C57Bl/6
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Large RNA samples were homogenized and extracted by Trizol; mRNAs were enriched through poly-A purification from a poly-A spin kit (NEB); small RNA samples were extracted using a miRvana microRNA isolation kit (samples 13-50) or total Trizol-extracted RNA (samples 51-93)
RNAseq library samples 1-6 were prepared using an Illumina TruSeq kit. RNAseq library samples 7-12 were prepared using a ScriptSeq v2 kit (Epicentre).
3ug of miRvana extracted or 5ug of Trizol extracted small RNAs were ligated to Linker-1 (IDT) in the absence of ATP with T4 RNA ligase 1 (New England Biolabs). Ligated RNA was size selected (based on an 18-36nt small RNA fraction) on 15% PAGE. The 5’ ends RNA were ligated to barcoded adapters. Dual-ligated RNA was reverse transcribed, PCR amplified and size selected on 4% NuSieve (Lonza) agarose. Small RNA sequencing was performed on the Illumina GAII (samples 13-50) and miSeq (samples 51-93) platforms.
|
| |
|
| Library strategy |
miRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina Genome Analyzer II |
| |
|
| Description |
total_shRNA_smallRNA_RPM.txt
|
| Data processing |
RNAseq reads were aligned to the mm9 genome through a standardized pipeline generated by the Stanford Center for Genomics and Personalized Medicine
FPKM values were calculated by use of Cufflinks v2.2.1 with the following parameters: “--library-norm-method quartile” and removing ribosomal, snoRNA and mitochondrial sequences.
small RNA sequencing reads were sorted by barcode, barcode removed (4nt), linker-1 clipped and aligned to mouse hairpins from mirBase release 15 using bowtie (version 0.12.7) with 2 mismatches and command -- best
small RNA sequences from 293 cells were aligned to the miR-122 plasmid that was transfected and raw counts for each small RNA that was generated were tabulated
Genome_build: mm9, mirBase release 15
Supplementary_files_format_and_content: File of FPKM values for RNAseq samples, of reads per million (RPM) mapped microRNA hits for mouse small RNA sequencing samples, and alignment to a transfected plasmid in human cell line samples, all in tab delimited text format
|
| |
|
| Submission date |
May 22, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Paul Valdmanis |
| E-mail(s) |
paulnv@uw.edu
|
| Phone |
206-221-8059
|
| Organization name |
University of Washington
|
| Department |
Medicine
|
| Lab |
Paul Valdmanis
|
| Street address |
1705 NE Pacific St, HSB J-309
|
| City |
Seattle |
| State/province |
WA |
| ZIP/Postal code |
98115-7720 |
| Country |
USA |
| |
|
| Platform ID |
GPL9250 |
| Series (1) |
| GSE69186 |
Small and large RNA sequencing of mouse livers receiving small hairpin RNAs |
|
| Relations |
| BioSample |
SAMN03732610 |
| SRA |
SRX1037301 |
