|
| Status |
Public on Jan 17, 2013 |
| Title |
GM0637 at T0 |
| Sample type |
RNA |
| |
|
| Source name |
GM0637 Fibroblast
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: Human Fibroblast
cell line: GM0637
treatment: none
|
| Biomaterial provider |
Coriell http://ccr.coriell.org/Sections/Search/Search.aspx?PgId=165&q=GM00637
|
| Treatment protocol |
GM0637 cells were treated with or without DNA damaging reagent neocarzinostatin (NCS), a radiomimetic drug that generates DSBs. Cells were harvested at varying time points (0-8 h).
|
| Growth protocol |
The GM0637 cells were plated on multiple 100-mm dishes for each line in 10% fetal calf serum and Dulbecco's modified Eagle's medium (DMEM) in 37℃ with 5% CO2.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol extraction of total RNA was performed according to the manufacturer's instructions (Invitrogen).
|
| Label |
Cy3
|
| Label protocol |
Sample labeling was performed according to the Agilent One-Color Microarray-Based Gene Expression Analysis protocol (Agilent Technology) with minor modifications. Briefly, mRNA was purified from 1 μg total RNA after removal of rRNA (mRNA-ONLY™ Eukaryotic mRNA Isolation Kit, Epicentre). Then, each sample was amplified and transcribed into fluorescent cRNA along the entire length of the transcripts without 3’ bias utilizing a random priming method. The labeled cRNAs were purified by RNAeasy Mini Kit (Qiagen). The concentration and specific activity of the labeled cRNAs (pmol Cy3/μg cRNA) were measured by NanoDrop ND-1000.
|
| |
|
| Hybridization protocol |
Array hybridization was performed according to the Agilent One-Color Microarray-Based Gene Expression Analysis protocol (Agilent Technology) with minor modifications. 1 μg of each labeled cRNA was fragmented by adding 11 μl 10 × Blocking Agent and 2.2 μl of 25 × Fragmentation Buffer, then heated the mixture at 60 °C for 30 min, finally 55 μl 2 × GE Hybridization buffer was added to dilute the labeled cRNA. 100 μl of hybridization solution was dispensed into the gasket slide and assembled to the LncRNA expression microarray slide. The slides were incubated for 17 hours at 65°C in an Agilent Hybridization Oven. The hybridized arrays were washed, fixed and scanned with using the Agilent DNA Microarray Scanner (part number G2505B).
|
| Scan protocol |
Agilent Feature Extraction software (version 10.5.1.1) was used to analyze acquired array images.
|
| Description |
Gene expression data from MCFs after DNA damage
|
| Data processing |
Quantile normalization and subsequent data processing were performed with using the GeneSpring GX v11.5.1 software package (Agilent Technologies). After quantile normalization of the raw data, LncRNAs and mRNAs that at least 1 out of 3 samples have flags in Present or Marginal (“All Targets Value”) were chosen for further data analysis.
|
| |
|
| Submission date |
Jan 15, 2013 |
| Last update date |
Jan 30, 2013 |
| Contact name |
Guohui Wan |
| E-mail(s) |
wanguohui1982@gmail.com
|
| Organization name |
The University of Texas M.D. Anderson Cancer Center
|
| Department |
Cancer Biology
|
| Lab |
SRB1.250
|
| Street address |
7777 Knight Road
|
| City |
Houston |
| State/province |
TX |
| ZIP/Postal code |
77054 |
| Country |
USA |
| |
|
| Platform ID |
GPL15314 |
| Series (1) |
| GSE43509 |
long noncoding RNAs expression data from GM0637 with and without DNA damage |
|
