|
| Status |
Public on Aug 05, 2015 |
| Title |
Clk-Mut CT34 |
| Sample type |
SRA |
| |
|
| Source name |
Liver
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57/BL6
tissue: Liver
genotype: Clock-delta-19, Scg2:tTA and tetO::Clock-HA
timepoint: CT34
|
| Treatment protocol |
3 days prior to constant conditions, ClockΔ19, Scg2:tTA and tetO::Clock-HA mice were treated with doxycycline in their drinking water (30 ug/ml). Treatment continued for the duration of the experiment.
|
| Growth protocol |
C57/BL6 mice with ClockΔ19 mutation as well as the Scg2:tTA and tetO::Clock-HA transgenes, were entrained to a 12h light : 12h dark schedule for one week. Mice were then transferred to complete darkness and housed in light-tight boxes for the duration of the experiment. Collection began one day following the shift to total darkness
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Liver chunks were dissected, snap frozen in liquid nitrogen, and RNA was extracted using Trizol reagent & RNeasy columns. RNA from 3 animals was pooled equally to create 40 ug of total RNA. PolyA RNA selected from total RNA using Invitrogen Dynabeads (cat# 610.06). Generate libraries using a modified version of Illumina Paired-End Library protocol (cat# PE-102-1001). RNA was fragmented for 5 minutes using Ambion Fragmentation Reagents (cat# AM8740). Fragmented RNA was converted into cDNA using Invitrogen SuperScript ds-cDNA kit (cat# 11917-010). Blunt ended cDNA fragments generated with Epicentre End Repair kit (cat# ER0720) and 'A' overhang created using NEB Klenow Fragment (cat# MO212s). Illumina Paired-End Adapters (from cat# PE-102-1001) ligated using Promega LigaFast kit (cat# M8221). Ligation products run on a 2% agarose gel and slice between 350-500bp cut from gel. Products purified using Qiagen Gel Extraction Kit (cat# 28704). Ligation products amplified with 13 PCR cycles using Illumina's paired-end primer mix (from cat# PE-102-1001) and NEB Phusion PCR Master Mix (cat# F-531S).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina Genome Analyzer IIx |
| |
|
| Data processing |
Align raw data using RNA-Seq Unified Mapper (RUM) to the mm9 genome, and RefSeq, UCSC, and Vega transcriptome annotations. RUM also calculated RPKM values for each transcript, exon, and intron in these annotations.
Genome_build: mm9
Supplementary_files_format_and_content: txt files contain RPKM values for each exon, intron, and transcript contained in the UCSC, RefSeq, and Vega annotations.
Supplementary_files_format_and_content: bedgraph files contain coverage plots for uniquely-aligned reads.
Supplementary_files_format_and_content: *.bed files contain high-confidence gapped-junctions mapped by uniquely-aligned reads.
|
| |
|
| Submission date |
Sep 21, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Nicholas Lahens |
| Organization name |
University of Pennsylvania
|
| Department |
ITMAT
|
| Street address |
Smilow Center for Translational Research 10-110 3400 Civic Center Blvd, Bldg 421
|
| City |
Philadelphia |
| State/province |
PA |
| ZIP/Postal code |
19104 |
| Country |
USA |
| |
|
| Platform ID |
GPL11002 |
| Series (1) |
| GSE41082 |
High-throughput sequencing of mouse liver transcriptome in Clock mutant animals |
|
| Relations |
| SRA |
SRX189135 |
| BioSample |
SAMN01180963 |
