|
| Status |
Public on Mar 20, 2014 |
| Title |
Stable FLAG-HA-CSB expression, biological replicate 1 |
| Sample type |
RNA |
| |
|
| Source name |
UVSS1KO cells stably transfected using linearized pFLAG-HA-CSB-IREShyg
|
| Organism |
Homo sapiens |
| Characteristics |
selection: hygromycin
immortalization: SV40
cell line: CSB-null UV-sensitive syndrome cells (UVSS1KO) cells
cell type: fibroblast
genotype/variation: CSB
|
| Treatment protocol |
pFLAG-HA-CSB-IREShyg3, pFLAG-HA-CSB-PGBD3-IREShyg3, and the empty vector pFLAG-HA-IREShyg3 were linearized with XhoI just downstream from the poly(A) site, and transfected using TransIT-LT1 reagent (Mirus) into UVSS1KO cells grown in DMEM. Selection with 100, 150 and 200 µg/ml hygromycin (Invitrogen) was begun after 24 h, and both drug and media were refreshed every 48–72 h. Confluent wells containing 50–100 colonies were trypsinized and passaged thereafter as pools. To generate doubly-transfected cells, the singly-transfected hygromycin-resistant pools were transfected with pFLAG-CSB-PGBD3-neo linearized by PvuI within the ampR gene. G418 selection was increased from 200 to 600 µg/ml while continuing 200 µg/ml hygromycin selection.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated by using RNeasy midi spin columns (Qiagen) and enriched for polyA+ RNA on Oligotex polydT beads (Qiagen).
|
| Label |
biotin
|
| Label protocol |
standard Affymetrix protocol
|
| |
|
| Hybridization protocol |
standard Affymetrix protocol
|
| Scan protocol |
standard Affymetrix protocol
|
| Description |
Gene expression data from CSB-null UVSS1KO cells expressing exogenous FLAG-HA-CSB to test the effects of CSB alone
UVSS1KO_CSB_1
|
| Data processing |
The 12 datasets were normalized using the Probe Logarithmic Intensity Error (PLIER) program of the Affymetrix Expression Console v1.1.1. using the Significance Analysis for Microarrays (SAM) program of the MeV v4.4 software suite.
|
| |
|
| Submission date |
Mar 20, 2014 |
| Last update date |
Mar 20, 2014 |
| Contact name |
Lucas T Gray |
| E-mail(s) |
[email protected]
|
| Organization name |
Allen Institute for Brain Science
|
| Department |
Molecular Genetics
|
| Lab |
Tasic
|
| Street address |
615 Westlake Ave N
|
| City |
Seattle |
| State/province |
WA |
| ZIP/Postal code |
98109 |
| Country |
USA |
| |
|
| Platform ID |
GPL570 |
| Series (1) |
| GSE56049 |
Transcriptional effects of CSB and the CSB-PGBD3 fusion protein in CSB-null UVSS1KO cells |
|
