|
| Status |
Public on Aug 01, 2007 |
| Title |
NHF3-hTERT_M_C3_26573 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Sample RNA
|
| Organism |
Homo sapiens |
| Characteristics |
Normal human fibroblast
|
| Biomaterial provider |
Kaufmann Lab
|
| Treatment protocol |
Colcemid at 100 ng/ml was added to G2 cells and cells were incubated an additional 4 h to collect cells in mitosis (M).
|
| Growth protocol |
Fibroblasts were cultured in DMEM high-glucose medium (Invitrogen, Carlsbad, CA, USA) supplemented with 2 mM L-glutamine (Invitrogen, Carlsbad, CA, USA) and 10% fetal bovine serum (FBS, Sigma Chemical Co, St. Louis, MO, USA)
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Qiagen RNeasy kit
|
| Label |
Cy3
|
| Label protocol |
1 µg of sample RNA was converted to cDNA with reverse transcriptase then amplified using T7 RNA polymerase while labeling with Cy3-dUTP (Agilent's Low RNA Input Linear Amplification Kit). The quality of each labeled cRNA was evaluated using an Agilent 2100 Bioanalyzer prior to hybridization.
|
| |
|
| Channel 2 |
| Source name |
Reference RNA
|
| Organism |
Homo sapiens |
| Characteristics |
global reference RNA
|
| Biomaterial provider |
Stratagene
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Stratagene protocol for preparation of reference RNA
|
| Label |
Cy5
|
| Label protocol |
1 µg of global reference RNA (Stratagene) was converted to cDNA with reverse transcriptase then amplified using T7 RNA polymerase while labeling with Cy5-dUTP (Agilent's Low RNA Input Linear Amplification Kit). The quality of each labeled cRNA was evaluated using an Agilent 2100 Bioanalyzer prior to hybridization.
|
| |
|
| |
| Hybridization protocol |
750ng of Cy3 and Cy5-labeled cRNA were used in the hybridization. The labeled cRNA from sham- or IR-treated samples was hybridized with the labeled global reference cRNA on an Agilent 22k human 1A array in a hybridization oven (Robbins Scientific Model 400, 1040-60-1AG) at 60oC for 17 h. Hybridization of sample RNA against reference RNA was done twice with dye swap.
|
| Scan protocol |
The arrays were scanned using the Agilent DNA Microarray Scanner with SureScan® Technology.
|
| Description |
The gene expression level was presented as ratio of sample intensity against reference intensity.
|
| Data processing |
Microarray images were analyzed using Agilent Feature Extraction Software (v7.1). Expression patterns and significant genes were extracted using Extracting Microarray Gene Expression Patterns and Identifying Biologically Significant Genes (EPIG)
|
| |
|
| Submission date |
Feb 19, 2007 |
| Last update date |
Feb 21, 2007 |
| Contact name |
Tong Zhou |
| E-mail(s) |
tong.zhou@gentris.com
|
| Phone |
919-724-9814
|
| Organization name |
Gentris Corperation
|
| Department |
Clincial Genetics
|
| Street address |
133 Southcenter Court, Suite 400
|
| City |
Morrisville |
| State/province |
NC |
| ZIP/Postal code |
27560 |
| Country |
USA |
| |
|
| Platform ID |
GPL887 |
| Series (1) |
| GSE7075 |
Identification of Primary Transcriptional Regulation of Cell-Cycle-Regulated Genes upon DNA Damage |
|
