|
| Status |
Public on Apr 01, 2018 |
| Title |
WT_1.cDNA.002.C74 |
| Sample type |
SRA |
| |
|
| Source name |
Human dermal fibroblasts
|
| Organism |
Homo sapiens |
| Characteristics |
disease: wild-type control
cell type: Fibroblast
passage number: P5
c1 chip size: Medium
|
| Growth protocol |
Dermal fibroblasts from patients with recessive dystrophic epidermolysis bullosa and their human leukocyte antigen (HLA) matched healthy siblings were obtained from skin biopsies and cultured in DMEM high glucose (Thermo Fisher Scientific) containing 10% fetal bovine serum (MilliporeSigma), 1% Pen/Strep (Thermo Fisher Scientific), 1% L-glutamine (Thermo Fisher Scientific), and 1% MEM NEAA (Thermo Fisher Scientific). For sub-culture, the medium was removed and cells were washed with 1X PBS (Thermo Fisher Scientific) and detached using Trypsin/EDTA (Thermo Fisher Scientific).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Fibroblasts were collected by trypsinization and resuspended in 5 ul of fibroblast medium for loading into the C1 capture chip.
cDNA was created from selected cells using the SMARTer Ultra Low RNA kit designed for the C1 system (Clontech). mRNA libraries were constructed using the the Nextera XT kit (Illumina) according to the manufacturer’s protocol.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina MiSeq |
| |
|
| Data processing |
Reads were mapped to hg19 using Bowtie2 (version 2.2.4) and Tophat (version 2.0.9)
Two single-cell samples sequenced as population controls with high read counts were sub-sampled (random sub-sampling, 10% of total reads)
Gene expression levels were calculated using Cuffquant (Cufflinks version 2.2.1 with parameters -u -max-bundle-frags 10000000) and Cuffnorm (Cufflinks version 2.2.1)
FPKM values as estimated by Cufflinks were added a value of 1 (to avoid zeros) and log2 transformed
Lowly expressed genes (average log2 (FPKM) < 1.5) were excluded from further analysis
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimted text file includes FPKM values
|
| |
|
| Submission date |
Jan 08, 2018 |
| Last update date |
Apr 01, 2018 |
| Contact name |
Jakub Tolar |
| E-mail(s) |
tolar003@umn.edu
|
| Organization name |
University of Minnesota, Twin Cities
|
| Department |
Pediatrics
|
| Street address |
420 Delaware St SE
|
| City |
Minneapolis |
| State/province |
MN |
| ZIP/Postal code |
55455 |
| Country |
USA |
| |
|
| Platform ID |
GPL15520 |
| Series (1) |
| GSE108849 |
Single-cell RNA-seq of fibroblasts from recessive dystrophic epidermolysis bullosa patients and wild-type controls |
|
| Relations |
| BioSample |
SAMN08325969 |
| SRA |
SRX3583426 |
