|
| Status |
Public on Sep 13, 2010 |
| Title |
WT cell, biological replicate 1 |
| Sample type |
RNA |
| |
|
| Source name |
ES cells expressing both Sall4a and Sall4b
|
| Organism |
Mus musculus |
| Characteristics |
cell line: CJ7
cell type: embryonic stem (ES) cells
sall4 isoform expression: Sall4a, Sall4b
|
| Treatment protocol |
Adherent cells were infected with concentrated lentivirus for 24 hrs in the presence of polybrene 2.5 mcg/mL. Media was then changed to ES cell media with LIF and puromycin (2 mcg/mL). Cells were then cultured for 48hrs post infection.
|
| Growth protocol |
CJ7 cells were grown under standard conditions prior to infection.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were lysed with Trizol and total RNA collected. Total RNA was further purified by the Promega S40 total rna isolation system per manufacturer's protocol. Total RNA was amplified by the Nugent process (http://chip.dfci.harvard.edu/index.php?option=com_content&task=view&id=14&Itemid=28#NuGen).
|
| Label |
biotin
|
| Label protocol |
The purified cDNA is fragmented through a chemical and enzymatic process. The fragmented product is labeled via enzymatic attachment of a biotin-labeled nucleotide to the 3’-hydroxyl end of the fragmented cDNA.
|
| |
|
| Hybridization protocol |
The biotinylated cDNA is added to a hybridization solution containing several biotinylated control oligonucleotides (for quality control), and hybridized to a microarray chip overnight at 45°C. The chips are then transferred to a fluidics instrument that performs washes to remove cDNA that has not hybridized to its complementary oligonucleotide probe. The bound cDNA is then fluorescently labeled using phycoerythrin-conjugated streptavidin (SAPE); additional fluors are then added using biotinylated anti-streptavidin antibody and additional SAPE.
|
| Scan protocol |
Each cDNA bound at its complementary oligonucleotide is excited using a confocal laser scanner, and the positions and intensities of the fluorescent emissions are captured. GeneChips were scanned using the Hewlett-Packard GeneArray Scanner G2500A.
|
| Description |
ES cells infected with a lentivirus with no shRNA.
WT1
|
| Data processing |
.gct and .res files were created by RMA normalization using GenePattern (https://www.broad.harvard.edu/cancer/software/genepattern/). These files (specifically the .gct) was used for downstream analysis by GSEA.
|
| |
|
| Submission date |
Mar 24, 2010 |
| Last update date |
Sep 13, 2010 |
| Contact name |
Sridhar Rao |
| E-mail(s) |
rao@bloodgroup.tch.harvard.edu
|
| Organization name |
Dana Farber Cancer Institute and Children's Hospital, Boston
|
| Department |
Pediatric Oncology
|
| Lab |
Karp Bldg, 7th floor
|
| Street address |
1 Blackfan Circle
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02115 |
| Country |
USA |
| |
|
| Platform ID |
GPL1261 |
| Series (2) |
| GSE21054 |
Differential roles of Sall4 isoforms in ES cell pluripotency: expression |
| GSE21056 |
Differential roles of Sall4 isoforms in ES cell pluripotency |
|
