 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Sep 24, 2018 |
| Title |
hESC2 |
| Sample type |
SRA |
| |
|
| Source name |
pluripotent stem cells
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Pluripotent cell
Stage: post-implantation stage
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The samples were processed directly using the SMARTer Ultra Low Input RNA for Illumina sequencing kit (Clontech Laboratories, Mountain View, CA, USA) to ensure minimum loss of cells. According to the manufacturer's suggested protocol, samples were collected in 2µl of DMEM-F12 + Glutamax (Invitrogen) with 25% bovine serum albumin (Sigma) and transferred to 5µl of pre-prepared reaction buffer. A final volume of 9µl was set up using nuclease-free water. The sample reaction mixture was then mixed with the first primer (3' SMART CDS primer II A) for first strand synthesis. This reaction mixture was placed in a thermocycler for 3 minutes and then incubated on ice. From this step, the first strand cDNA synthesis was set up directly using the SMARTScribe Reverse Transcriptase. Once completed, the first-strand cDNA was purified using SPRI Ampure beads, followed by double stranded (ds) cDNA amplification by long distance PCR. The amplified ds cDNA was then further purified using the Ampure beads.
cDNA was fragmented in the 200-500bp range on a Covaris S2 Focused-ultrasonicator (Brighton, UK), according to the SMARTer Ultra Low Input RNA for Illumina sequencing protocol. A sequencing library was made using the Low Input Library Prep Kit (Clontech Laboratories, Mountain View, CA, USA).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
Primed hESCs (XX) were maintained in feeder-free culture conditions on Matrigel for two passages and approximately 50 cells were collected
|
| Data processing |
Reads were trimmed using cutadapt versin 1.11 to remove the Illumina adaptor sequences
The reads were mapped against the Homo sapiens GRCh37.74 reference genome using CLC genomics workbench version 9.0.1 (CLC)
Count tables were made by counting the reads that mapped against the genes defined by the Ensembl GRCh37.74 Gene Transfer Format file (GTF) and exported from CLC
Genome_build: Homo sapines GRCh37.74
Supplementary_files_format_and_content: comma separated values files including Gene counts, Gene ID and other gene information
|
| |
|
| Submission date |
Sep 03, 2018 |
| Last update date |
Sep 24, 2018 |
| Contact name |
Laurentijn Tilleman |
| Organization name |
Ghent University
|
| Street address |
Ottergemsesteenweg 460
|
| City |
Gent |
| ZIP/Postal code |
9000 |
| Country |
Belgium |
| |
|
| Platform ID |
GPL18573 |
| Series (1) |
| GSE119378 |
Transcriptional landscape changes during human embryonic stem cell derivation |
|
| Relations |
| BioSample |
SAMN09948066 |
| SRA |
SRX4634280 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3373933_hESC2.csv.gz |
2.7 Mb |
(ftp)(http) |
CSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
|
|
|
|
 |
