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Status |
Public on May 29, 2014 |
Title |
e9.5_TGC_female_3seq_rep2 |
Sample type |
SRA |
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Source name |
Trophoblast giant cells - e9.5
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Organism |
Mus musculus |
Characteristics |
strain: C57BL/6 cell type: e9.5 trophoblast giant cells
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Growth protocol |
Pregnant C57BL/6 mice were obtained from Charles River. Copulation was determined by the presence of a vaginal plug the morning after mating, and embryonic day 0.5 (e0.5) was defined as noon of that day. TGCs and embryos were dissected in 1X PBS (1:10 10X PBS, pH=7.4; Gibco) and stored on ice until further processing. TGCs were dissected away from the placental disk, decidua, and, if possible, Reichert’s membrane. For gathering RNA, the entire embryo was collected. All experimental procedures were carried out in accordance with the APLAC protocol and the institutional guidelines set by the Veterinary Service Center at Stanford University.
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Extracted molecule |
polyA RNA |
Extraction protocol |
mRNA was extracted directly from cell lysates using Dynabeads Oligo (dT)25 (Invitrogen). 500 ng of mRNA was then used to prepare 3’ RNA-Seq libraries as previously described (Beck et al, PLOS One 2010). Briefly, mRNA was heat sheared for 7.5 minutes to produce an average fragment size range of 100-400 bp, then used to generate cDNA libraries using a custom oligo dT primer containing Illumina-compatible adapter sequence. cDNA fragments were end-repaired and ligated to standard Illumina adapters. Size-selection was performed using E-gel SizeSelect agarose gels (Invitrogen), products were PCR amplified for 15 cycles, and purified using Ampure XP beads. Library quality was assessed using Bioanalyzer and Qubit, and sequenced on the Genome Analyzer IIx.
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Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina Genome Analyzer IIx |
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Description |
Tissue 3' RNA-Seq
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Data processing |
Basecalls performed using CASAVA version 1.5 3' RNA-seq reads were aligned to mm9 or rn4 using Bowtie 0.12.7 with the settings --best --strata -k 10 -S --chunkmbs 190 Using Unipeak v1.0, Bowtie alignments were filtered by posterior probability (sam_map.pl from Unipeak), and transcription peaks were called using default settings. Peaks (chromosomal coordinates) were associated with Ensembl (e67) gene database for mouse and multiple peaks for each gene were pooled. Read counts were normalized across libraries using DESeq genome build: = mm9 supplementary files format and content: txt files contain pooled gene x normalized tag count
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Submission date |
Sep 03, 2013 |
Last update date |
May 15, 2019 |
Contact name |
Roberta L Hannibal |
E-mail(s) |
robertah@stanford.edu
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Organization name |
Stanford University
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Department |
Genetics
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Street address |
300 Pasteur Drive
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City |
Stanford |
State/province |
CA |
ZIP/Postal code |
94305 |
Country |
USA |
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Platform ID |
GPL11002 |
Series (2) |
GSE50561 |
Trophoblast giant cell underrepresentation (sequencing) |
GSE50585 |
Trophoblast giant cell underrepresentation |
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Relations |
Reanalyzed by |
GSE80797 |
BioSample |
SAMN02344599 |
SRA |
SRX344636 |
Supplementary data files not provided |
SRA Run Selector |
Raw data are available in SRA |
Processed data are available on Series record |
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