|
| Status |
Public on Dec 24, 2013 |
| Title |
HEK293-EGFP-ZFP36_ZFP36_Rep2 |
| Sample type |
RNA |
| |
|
| Source name |
FlpIn T-Rex Human embryonic kidney 293 stable cell line
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: HEK293
cell: EGPF-ZFP36_Stable
treatment: Doxycline
|
| Treatment protocol |
Expression of stable transgenes were induced by incubation with 1 µg/ml doxycyline for 16 h.
|
| Growth protocol |
Doxycycline-inducible epitope-tagged ZFP36 HEK293 cell lines were produced utilizing the Flp-In™ T-REx™ 293 Cell Line system (Life Technologies). Flp-In™ T-REx™ HEK293 cells were generated that contained a FLAG/HA-tagged human ZFP36 or an EGFP-tagged human ZFP36. Both cell lines were cultured in DMEM supplemented with 10% tet-reduced FBS and appropriate selection antibiotics. For induction of FLAG/HA-tagged or EGFP-tagged ZFP36 protein, cells were treated for 16 hrs with 1 μg/ml doxycycline (Sigma).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total or immunoprecipitated RNA was extracted by phenol-chloroform, followed by ethanol precipitation. Rneasy Mini Kit (Invitrogen) was used according to manufacturer's instructions as a final step.
|
| Label |
biotin
|
| Label protocol |
1.5 μg of RNA from each sample was provided to the NIEHS Microarray Core Facility for analysis, using Affymetrix Human Genome U133 Plus 2.0 gene chip (Affymetrix, Santa Clara, CA). One hundred ng of total RNA was amplified as directed in the Affymetrix 3' IVT Express kit protocol.
|
| |
|
| Hybridization protocol |
12.5 μg of amplified biotin-aRNAs were fragmented and 10 μg were hybridized to each array for 16 hours at 45° C in a rotating hybridization oven using the Affymetrix Eukaryotic Target Hybridization Controls and protocol.
|
| Scan protocol |
Arrays were scanned in an Affymetrix Scanner 3000 and data was obtained using the GeneChip® Command Console Software (AGCC; Version 1.1)
|
| Data processing |
Data was normalized using gcrma and then filtered for retaining only unambiguously mapping probes and minimal expression level (mean log hybridization value for replicates within a group > 4.5).
|
| |
|
| Submission date |
Dec 10, 2013 |
| Last update date |
Dec 25, 2013 |
| Contact name |
Neelanjan Mukherjee |
| E-mail(s) |
[email protected], [email protected]
|
| Phone |
3037241623
|
| Organization name |
University of Colorado School of Medicine
|
| Department |
BMG/RBI
|
| Lab |
Neelanjan Mukherjee
|
| Street address |
12801 E 17th Ave
|
| City |
Aurora |
| State/province |
CO |
| ZIP/Postal code |
80045 |
| Country |
USA |
| |
|
| Platform ID |
GPL570 |
| Series (2) |
| GSE53183 |
Global Target mRNA Specification and Regulation by RNA-Binding Protein ZFP36/Tristetraprolin/TTP [Affymetrix] |
| GSE53185 |
Global Target mRNA Specification and Regulation by RNA-Binding Protein ZFP36/Tristetraprolin/TTP |
|
