|
| Status |
Public on May 31, 2020 |
| Title |
MDA-MB-231-205-5p Rep1 |
| Sample type |
RNA |
| |
|
| Source name |
MDA-MB-231-205-5p
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: MDA-MB-231
cell type: Breast cancer cell line
miRNA: has-miR-205-5p precursor
|
| Treatment protocol |
MDA-MB-231 cells (0.168 x 10e6 cells/ml) were transfected with the selected miRNA precursors (pre-miR–negative control, hsa-miR-205-5p and hsa-miR-214-3p) purchased from Ambion. Cell transfection was conducted employing a reverse transfection protocol.
|
| Growth protocol |
MDA-MB-231 cells were grown in DMEM supplemented with 10% fetal bovine serum, 1% NEAA, 1% L-glutamine, 100 mg/l penicillin, and 100 mg/l streptomycin.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA were isolated from control and treated cells using Total RNA Purification Kit (Norgen-Biotek Corp., Canada) according to the manufacturer’s instructions. The concentrations of total RNA were measured using NanoDrop 2000 (Thermo-Scientific).
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 100 ng RNA using the Low Input Quick Amp Labeling Kit, one-color (Agilent) according to the manufacturer's instructions, followed by RNeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-2000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
1.65 ug of Cy3-labelled cRNA was fragmented at 60°C for 30 minutes in a reaction volume of 55 ul following the manufacturer’s instructions. On completion of the fragmentation reaction, 55 ul of 2× Hi-RPM Hybridization Buffer (Agilent) was added to the fragmentation mixture and hybridized to Agilent Whole Human Genome GE 4x44K v2 Microarray chip 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.
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505C) using one color scan setting for 8x60k array slides.
|
| Description |
Gene expression in MDA-MB-231 breast cancer cell line transfected with has-miR-205-5p precursor replica 1.
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 11.0 (Agilent) using default parameters. Data were analyzed using Agilent GeneSpring X software.
|
| |
|
| Submission date |
Apr 17, 2020 |
| Last update date |
Jun 01, 2020 |
| Contact name |
Nehad M Alajez |
| E-mail(s) |
[email protected]
|
| Organization name |
King Saud University College of Medicine
|
| Department |
Anatomy
|
| Lab |
Stem Cell Unit
|
| Street address |
1st floor College of Medicne
|
| City |
Riyadh |
| ZIP/Postal code |
11461 |
| Country |
Saudi Arabia |
| |
|
| Platform ID |
GPL17077 |
| Series (2) |
| GSE148847 |
Gene expression profiling of MDA-MB-231 cells transfected with hsa-miR-205-5p and hsa-miR-214-3p compared to scrambled miRNA precursors |
| GSE148848 |
microRNA expression profiling of paired primary and lymph node metastatic breast cancer patients, and gene expression profiling of MDA-MB-231 cells transfected with hsa-miR-205-5p and hsa-miR-214-3p compared to scrambled miRNA precursors |
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