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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Oct 03, 2012 |
| Title |
Mouse_TG_GD_RNA-seq |
| Sample type |
SRA |
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| Source name |
pool of thyroid gland tissues from group
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| Organism |
Mus musculus |
| Characteristics |
tissue: thyroid gland
generation: F1H (F1, first generation, H hybrid)
strain: hybrid C57BL/6xBALB/c
genotype/variation: CD40 over-expressing transgenic
treatment protocol: immunized with thyrotropin receptor A subunit cDNA in an adenovirus construct
immunization outcome: developed EAGD
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| Treatment protocol |
Primary thyroid cell cultures were washed with 10ml PBS after 24 hours, and then 10mL of medium [DMEM, 4.0 mM L-glutamine and sodium pyruvate, 10% FBS, 1% antibiotic anti-mycotic solution (all from Hyclone/Thermo Fisher Waltham, MA)] were added to the flask. This was allowed to incubate at 37oC with 5% CO2 overnight. Cell were washed with PBS, and plated evenly in a 12 well plate. Cells were incubated 24 hours to allow for adherence to the wells. Again, the cells were washed and at this time the cells were treated for 0 or 24 hours with 1 ug of anti-CD40 stimulating antibody (G28.5).
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| Growth protocol |
Human thyroid primary cells were prepared from fresh, non-cancerous thyroid tissue adjacent to thyroid tumors that were removed at surgery. Tissue was minced and digested in a 50 ml falcon tube with type II collagenase at a concentration of 5 mg/ml for three hours at 37 degrees celsius. After three hours, 10ml of medium [DMEM, 4.0 mM L-glutamine and sodium pyruvate, 10% FBS, 1% antibiotic anti-mycotic solution was added to the tube, and the product was strained through a 70 um cell strainer into another 50 ml falcon tube. The tube was centrifuged at 1000 rpm for 5 minutes. The supernatant was carefully removed from the pellet and the pellet was washed with another 10 ml of medium. Again, the tube was spun at 1000 rpm for 5 minutes. The medium was removed from the pellet. The pellet was suspended in 10 ml of medium, and the cells were plated in a 25 cm2 flask, and placed in an incubator at 37 degrees celsius with 5% CO2.
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| Extracted molecule |
total RNA |
| Extraction protocol |
According to Illumina mRNA Sequencing Sample Preparation Guide. Briefly, 2ug total RNA hybridized to poly dT magnetic beads to capture polyA mRNA molecules. Chemical fragmentation, first strand synthesis with RTase, randomers second strand synthesis. Then end repair, 3' A overhang, multiplex adapter ligation (from Illumina indexing kit), size selection by electrophoresis and cut at 200bp. 18 cycles of PCR with a primer bearing an indexing barcode.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
RNA seq of thyroid gland tissues of transgenic F1H mice immunized that did develop EAGD
mouse_TGEAGD
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| Data processing |
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to human hg18 or mouse mm9 exon and exon junction sequences and contamination datanases using bwa algorithm
After filtering reads mapped to contamination databases, the reads that have one or no mismatch and are uniquely aligned to each exon and splicing-junction sites were extracted and then counted. The read count for each RefSeq transcript was calculated by combining the counts for exons and splicing junctions of corresponding transcript.
Supplementary_files_format_and_content: tab-delimited txt file including count values for RefSeq transcripts
Genome_build: hg18 or mm9
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| Submission date |
Jul 03, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Weijia Zhang |
| E-mail(s) |
weijia.zhang@mssm.edu
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| Phone |
212-241-2883
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| Organization name |
Mount Sinai School of Medicine
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| Department |
Department of Medicine
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| Lab |
Bioinformatics Lab
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| Street address |
1425 Madison Avenue
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| City |
New York |
| State/province |
NY |
| ZIP/Postal code |
10029 |
| Country |
USA |
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| Platform ID |
GPL13112 |
| Series (1) |
| GSE39081 |
Genetically-driven target tissue over-expression of CD40: A novel mechanism in autoimmune disease |
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| Relations |
| SRA |
SRX157316 |
| BioSample |
SAMN01085222 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM955429_Mouse_TG_GD_RefSeq_count.txt.gz |
192.8 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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