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| Status |
Public on Dec 10, 2014 |
| Title |
M7838_uninvolved |
| Sample type |
SRA |
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| Source name |
skin
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| Organism |
Homo sapiens |
| Characteristics |
tissue type: Uninvolved skin
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| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was prepared from the frozen biopsies using the Qiagen Allprep kit. Skin punch biopsies were trimmed of visible subcutaneous fat and visible blood was removed by blotting on saline-soaked gauze. Biopsies were then snap frozen in liquid nitrogen and stored at -80 deg C. For RNA isolation, frozen skin biopsy samples were wrapped in an aluminum foil pouch pre-frozen in liquid nitrogen, placed in a prechilled steel mortar, and crushed with a hammer. The pulverized frozen tissue was quickly transferred into a 1.5-ml microcentrifuge containing 1 ml of the Allprep buffer and glass grinding beads. The tube was vigorously agitated on a Vortex mixer for 5 minutes to complete the extraction process. From this point on the sample was processed exactly as described in the kit manual. RNA quality and quantity were checked using an Agilent Bioanalyzer.
Libraries for high throughput sequencing were prepared using the Illumina mRNA-Seq kit. Briefly, the process involved isolating polyadenylated RNA, fragmentation by heating at 90°C, random-primed cDNA preparation, ligation of adaptors, and size selection on an agarose gel. A gel slice corresponding to approximately 120 bp insert was isolated, and the DNA was extracted and amplified by 15 cycles of PCR using the adaptor primers. The gel purified final product was checked for quality and quantity on an Agilent Bioanalyzer. The library thus prepared was sequenced one sample per lane in an 8-lane flow cell on the Illumina Genome Analyzer IIx. The raw sequencing output of the 80-base single read sequencing was 2000-3000 Mb per lane corresponding to 25-37.5 million reads.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina Genome Analyzer |
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| Data processing |
Sequenced reads were aligned to the reference genome NCBI build 37 using TopHat version 1.3.3. We only used uniquely mapped reads with less than 3 mismatches.
We constructed a dataset-dependent transcriptome (see the linked article for detail), and the expression level of each gene was obtained by counting the number of reads. Reads mapped to multiple exons in a gene were counted only once. The gtf file showed the chromosome coordinates for each gene identifier in the ReadCount_42samples file.
Genome_build: genome NCBI build 37
Supplementary_files_format_and_content: data matrix (ReadCount_42samples.txt) containing read counts and gtf file showing the chromosome coordinates for each gene identifier in the ReadCount_42samples.txt file
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| Submission date |
Dec 09, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Lam Tsoi |
| E-mail(s) |
[email protected]
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| Organization name |
University of Michigan
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| Street address |
Medical Sciences 1 Bldg. 1301 E. Catherine
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| City |
Ann Arbor |
| State/province |
MI |
| ZIP/Postal code |
48109 |
| Country |
USA |
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| Platform ID |
GPL9052 |
| Series (2) |
| GSE63979 |
Analysis of long non-coding RNAs highlights tissue-specific expression patterns and epigenetic profiles in normal and psoriatic skin |
| GSE63980 |
Transcriptomic studies to characterize the gene expression landscape of psoriasis |
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| Relations |
| BioSample |
SAMN03256287 |
| SRA |
SRX800855 |
| 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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