|
| Status |
Public on Apr 05, 2024 |
| Title |
UNTREATED HCT116 COLORECTAL CANCER CELLS REPLICATE 4 |
| Sample type |
RNA |
| |
|
| Source name |
control group
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: colorectal cancer cell line
gender: male
treatment: untreated
|
| Treatment protocol |
Cells were treated 2.5 mM Metformin for two weeks. The DMEM media containing Metformin was changed every day.
|
| Growth protocol |
HCT116 cells were grown in 5% CO2 at 37°C. Cells were grown in DMEM media and subcultured every third day using trypsin-EDTA.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was prepared using Trizol reagent. The protocol includes brief pipetting and vorteing, followed by phase seperation by chloroform and precipitation using iso-propanol. This was followed by washing by 75% ethanol and elution in nuclease-free waster. Total RNA was quantified using a NanoDrop-2000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer.
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 200 ng total RNA using the One-Color Low TOTAL RNA Input Linear Amplification kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification. Dye incorporation and cRNA yield were checked with the NanoDrop ND-2000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
0.83 ug of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 250 ml containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 250 ml of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole Human Genome Oligo Microarrays for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation.
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505B) using one color scan setting for 8x60k array slides (Scan Area 61x21.6 mm, Scan resolution 5 um, Dye channel is set to Green and Green PMT is set to 100%).
|
| Description |
GE2207_2_4
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.7.3.1 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
|
| |
|
| Submission date |
Apr 05, 2021 |
| Last update date |
Apr 05, 2024 |
| Contact name |
Muzaffer Ahmed Bhat |
| E-mail(s) |
muzaffar.sciml87@gmail.com
|
| Phone |
91 8826753204
|
| Organization name |
AIIMS
|
| Department |
Physiology
|
| Lab |
Molecular Biology
|
| Street address |
ANSARI NAGAR NEW DELHI 110029
|
| City |
Delhi |
| State/province |
Delhi |
| ZIP/Postal code |
110029 |
| Country |
India |
| |
|
| Platform ID |
GPL14550 |
| Series (1) |
| GSE171478 |
HCT116 colorectal cancer cells treated with 2.5 mM Metformin |
|
