|
| Status |
Public on Apr 04, 2007 |
| Title |
TGF-beta treated lung epithelial cells at 12hr_Exp2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
lung epithelial cells control
|
| Organism |
Homo sapiens |
| Characteristics |
Lung epithelial cell-line immortalized with SV-40 large T antigen
|
| Biomaterial provider |
T. Takahashi
|
| Treatment protocol |
No treatment (Control for 5ng/ml TGF-beta1treatment), grown in recommended medium, washed with serum free medium and allowed to remain in serum free medium for 12hr before extraction of the sample.
|
| Growth protocol |
Cells were plated in tissue culture dishes in Ham’s F-12 medium with 5% Fetal Bovine Serum and other constituents, incubated at 37 degrees C in 5% CO2 and 90% humidity.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was Extracted using TRI reagent from Sigma Aldrich with an extra chloroform extraction and cleaned up using RNA easy mini columns.RNA was quantitated on a spectrophotometer and analyzed on a formaldehyde-agarose gel.
|
| Label |
Cy3
|
| Label protocol |
The labelling of the probe was done using the Micromax direct labelling kit (Perkin Elmer Life Sci.) according to the recommended protocol.
|
| |
|
| Channel 2 |
| Source name |
lung epithelial cells TGF-beta treated
|
| Organism |
Homo sapiens |
| Characteristics |
Lung epithelial cell-line immortalized with SV-40 large T antigen
|
| Biomaterial provider |
T. Takahashi
|
| Treatment protocol |
Cells grown in recommended medium, washed with serum free medium and treated with 5ng/ml TGF-beta1 in serum free medium, for 12hr before extraction of the sample.
|
| Growth protocol |
Cells were plated in tissue culture dishes in Ham’s F-12 medium with 5% Fetal Bovine Serum and other constituents, incubated at 37 degrees C in 5% CO2 and 90% humidity.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was Extracted using TRI reagent from Sigma Aldrich with an extra chloroform extraction and cleaned up using RNA easy mini columns.RNA was quantitated on a spectrophotometer and analyzed on a formaldehyde-agarose gel.
|
| Label |
Cy5
|
| Label protocol |
The labelling of the probe was done using the Micromax direct labelling kit (Perkin Elmer Life Sci.) according to the recommended protocol.
|
| |
|
| |
| Hybridization protocol |
The hybridisation was carried out in the GeneTAC Hyb Station (Genomic solutions, UK) at 65 degree for 4 hours, 60 degree for 4 hours and 55 degree for 10 hours. The slides were then washed with medium stringency (2X SSC and 0.1% SDS), high stringency (0.1X SSC and 0.1% SDS) and post wash (0.1X SSC) buffers for 5 minutes each, dried and scanned using a Scanner (Scanarray Express, Perkin Elmer life Sciences, USA).
|
| Scan protocol |
Scanned using Scanarray Express (Perkin Elmer Life Sci), the software used for image aquisition was scanarray
|
| Description |
no additional information is necessary
|
| Data processing |
All the image analyses have been done using the Quantarray software (Perkin Elmer Life Sciences, USA). Filtering and compilation of data have been done using Microsoft Excel and Microsoft Access. Spots of compromised quality and with low intensity were eliminated from the analysis. The data was normalized by LOWESS method (Avadis 3.1, Strand Life Sciences, India), Cy5:Cy3 ratios were established and log2 values were calculated.
|
| |
|
| Submission date |
Apr 03, 2007 |
| Last update date |
Apr 04, 2007 |
| Contact name |
Paturu Kondaiah |
| E-mail(s) |
[email protected]
|
| Phone |
91-80-22932688
|
| Fax |
91-80-23600999
|
| Organization name |
Indian Institute of Science
|
| Department |
Molecular Reproduction, Development and Genetics
|
| Street address |
C.N.R. Rao Circle
|
| City |
Bangalore |
| ZIP/Postal code |
560012 |
| Country |
India |
| |
|
| Platform ID |
GPL3515 |
| Series (1) |
| GSE7436 |
Profiling of genes regulated by TGF-beta in lung carcinoma (A549) and immortalized lung epithelial (HPL1D)cells. |
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