|
| Status |
Public on Apr 12, 2017 |
| Title |
siGRHL3 replicate 1, 12 hours migration, NHEK [siGRHL3-migration] |
| Sample type |
RNA |
| |
|
| Source name |
NHEK
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: migrating
cell line: NHEK
|
| Treatment protocol |
100,000 cells were plated in each well of a 12 well plate, and were transfected with 30nM control (Dharmacon D-001810-10-05), or GRHL3 (Dharmacon L-014017-02). To induce migration, cells were grown to a confluent monolayer and scratched with a p200 pipette tip. Scratches were made approximately every 1 cm horizontally and vertically, resulting in a grid of scratches. Migration was induced 60 hours after transfection, and cells were collected 12 hours later (72 hours after transfection).
|
| Growth protocol |
Normal Human Epidermal Keratinocytes (NHEK) were purchased from LifeLine Technologies and grown according to the manufacturer’s instructions in DermaLife medium (LifeLine Tech) supplemented with DermaLife growth factors (LifeLine Tech).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were collected and lysed in Trizol, followed purification with Zymo RNA extraction kit. RNA concentration and quality were quantified on a NanoDrop.
|
| Label |
biotin
|
| Label protocol |
The Ambion WT expression kit (Life Technologies) was used to prepare RNA samples for whole transcriptome microarray analysis
|
| |
|
| Hybridization protocol |
2ug of the labeled, fragmented single stranded cDNA is hybridized at 45C with rotation for 17 hours (Affymetrix GenechIP Hybridization Oven 640) to probe sets present on an Affymetrix GeneChIP 1.0ST array. The GeneChIP arrays were washed and then stained with streptavidin-phycoerythrin on an Affymetrix Fluidics Station 450 (Fluidics protocol FS450_007)
|
| Scan protocol |
Arrays were scanned using GeneChIP Scanner 3000 7G and Command Console Software v. 3.2.3 to produce .CEL intensity files.
|
| Description |
Grhl3-12-1
|
| Data processing |
The probe cell intensity files (*.CEL) were analyzed in Affymetrix Expression Console software v1.1.1 using the PLIER algorithm to generate probe level summarization files (*.CHIP). The settings used were algorithm-PLIER v2.0; quantification scale-Linear;quantification type-signal and detection p-value; background-PM-GCBG; normalization method-sketch-quantile.
Raw values lower than 200 were not considered expressed
|
| |
|
| Submission date |
Feb 03, 2017 |
| Last update date |
Apr 13, 2017 |
| Contact name |
Bogi Andersen |
| Organization name |
University of California, Irvine
|
| Department |
Medicine
|
| Lab |
Andersen
|
| Street address |
839 Health Sciences Dr.
|
| City |
Irvine |
| State/province |
CA |
| ZIP/Postal code |
92697 |
| Country |
USA |
| |
|
| Platform ID |
GPL6244 |
| Series (2) |
| GSE94465 |
GRHL3 binding and the enhancer landscape are reorganized during transitions between different functional states of epidermal keratinocytes [siGRHL3-migration] |
| GSE94471 |
GRHL3 binding and the enhancer landscape are reorganized during transitions between different functional states of epidermal keratinocytes |
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