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| Status |
Public on Aug 12, 2020 |
| Title |
HUVEC AD-HIES2_TNF |
| Sample type |
SRA |
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| Source name |
Umbilical Cord
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| Organism |
Homo sapiens |
| Characteristics |
isolated cells: HUVEC
treatment: TNFa 50ng/ml for 8h
genotype/variation: AD-HIES STAT3 mutation
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| Treatment protocol |
For RNA isolation, HUVECs were seeded on 6 well plates at density of 250,000 cells per well. In 48 hours, when cells reached confluence, media was changed to fresh one with or without 50ng/ml TNFα for 8h and total RNA was extracted for RNA-Seq analysis.
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| Growth protocol |
The cords were shipped overnight from hospitals in PBS supplemented with Penicillin-Streptomycin (No. 15140122, ThermoFisher Scientific). Umbilical veins were washed 3 times with PBS supplemented with Penicillin-Streptomycin and Amphotericin B (No. 400-104, Gemini Bio-Products, West Sacramento, CA) and then filled with collagenase, type II (No.17101-015, ThermoFisher Scientific) at 0.2% in PBS and the ends were closed. The cords were placed in 37C incubator for 20 min. The cells were pelleted from the collagenase suspension, resuspended in endothelial cell culture media (EBM Basal Medium (No.CC-3121, Lonza) supplemented with EGM-2 SingleQuots Kit (No.CC-4176, Lonza)), Penicillin-Streptomycin and Amphotericin B and plated on fibronectin (No. F2006, Sigma) coated T25 flask. The media was changed next day, and cells were routinely cultured on 10 cm fibronectin coated dishes.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using TRIzol Reagent (No. 15596026, ThermoFisher Scientific).
One mg of total RNA was used as input for library preparation using the TruSeqRNA Sample Preparation Kit v2 (Illumina) according to manufacturer’s instructions. Sequencing libraries were quantitated by qPCR on a LightCycler 480 Instrument II (Roche) and assessed for size distribution and absence of adapter dimers using a Fragment Analyzer (AATI). Libraries were pooled for multiplexing and sequencing was performed on the Illumina HiSeq 3000 Sequencing System with run parameters at paired-end 75bp read length for >25 million paired-end reads per sample.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 3000 |
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| Description |
HUVEC_RNA-seq.txt
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| Data processing |
RNA-seq data quality was assessed using FastQC
Reads obtained for each sample were aligned to the hg19 human reference genome using Tophat2
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimited text file include total counts values for all samples
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| Submission date |
Oct 21, 2019 |
| Last update date |
Aug 12, 2020 |
| Contact name |
Manfred Boehm |
| Organization name |
NHLBI
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| Street address |
9000 Rockbille Pike
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| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
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| Platform ID |
GPL21290 |
| Series (2) |
| GSE139188 |
Gene expression analysis in endothelial cells derived from patients with Autosomal-dominant hyper-IgE syndrome (AD-HIES) |
| GSE139365 |
Gene expression analysis in primary cells derived from patients with Autosomal-dominant hyper-IgE syndrome (AD-HIES) |
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| Relations |
| BioSample |
SAMN13071237 |
| Supplementary data files not provided |
| Raw data not provided for this record |
| Processed data are available on Series record |
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