|
| Status |
Public on Sep 24, 2022 |
| Title |
Rat dermal fibroblast, 48h, control lentivirus, replicate1 |
| Sample type |
RNA |
| |
|
| Source name |
Rat dermal fibroblast, 48h, control lentivirus
|
| Organism |
Rattus norvegicus |
| Characteristics |
cell type: fibroblast
treatment: control lentivirus
genotype: control
|
| Treatment protocol |
A lnc-URIDS shRNA lentivirus and a control lentivirus (GenePharma, Shanghai, China) were transfected into rat dermal fibroblasts at a multiplicity of infection (MOI) of 10, and 2 µg/mL puromycin (Thermo Fisher Scientific) for 48h was added to the medium of the cells.
|
| Growth protocol |
Dermal fibroblasts were cultured in Dulbecco’s modified Eagle’s medium (DMEM) containing 1,000 mg/L glucose (Gibco, Gaithersburg, MD) and 10%FBS.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from the cells using TRIzol reagent (AGbio, China) following the manufacturer’s instructions.RNA quantity and quality were measured by NanoDrop ND-1000. RNA integrity was assessed by standard denaturing agarose gel electrophoresis.
|
| Label |
Cy3
|
| Label protocol |
Sample labeling were performed according to the Agilent One-Color Microarray-Based Gene Expression Analysis protocol (Agilent Technology). Briefly, total RNA from each sample was linearly amplified and labeled with Cy3-UTP. The labeled cRNAs were purified by RNeasy 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 were performed according to the Agilent One-Color Microarray-Based Gene Expression Analysis protocol (Agilent Technology). 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 gene expression microarray slide. The slides were incubated for 17 hours at 65°C in an Agilent Hybridization Oven.
|
| Scan protocol |
The hybridized arrays were washed, fixed and scanned with using the Agilent DNA Microarray Scanner (part number G2505C).
|
| Description |
Gene expression after 48hr trasfected control lentivirus
|
| Data processing |
the scanned images were analysed using Agilent Feature Extraction software (v11.0.0.1).
Normalization of signal intensity using Agligent GeneSpring GX v12.1 software
|
| |
|
| Submission date |
Sep 23, 2022 |
| Last update date |
Sep 24, 2022 |
| Contact name |
Meng Ren |
| E-mail(s) |
[email protected]
|
| Phone |
+862081332286
|
| Organization name |
Sun Yat-sen Memorial Hospital, Sun Yat-sen University
|
| Department |
endocrinology department
|
| Street address |
107 Yanjiang West Road
|
| City |
guangzhou |
| ZIP/Postal code |
510120 |
| Country |
China |
| |
|
| Platform ID |
GPL14746 |
| Series (1) |
| GSE214034 |
Development of gene expression signatures for lnc-URIDS knock down rat dermal fibroblast |
|
