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| Status |
Public on Aug 31, 2018 |
| Title |
PSC27-VCR-treated-rep 2 |
| Sample type |
RNA |
| |
|
| Source name |
PSC27 VCR-treated, 100 nM vincristine for 12 hrs, replicate 2
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Normal human prostate
cell type: Primary stromal cells
|
| Treatment protocol |
For experimental groups, cells were either treated with either ionizing radiation (Cs137) for 10Gy, 50 ug/ml bleomycin for 6 hours, 1 uM mitoxantrone for 24 hours, 5 uM docetaxel for 12 hours, 1 uM paclitaxel for 12 hours, or 100 nM vincristine for 12 hours.
|
| Growth protocol |
PSC27 cells were grown in prostate stromal cell complete (PSCC) media supplemented with 10% FBS and 1% antibiotics.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNAs were prepared with a RNeasy Miniprep kit (Qiagen) according to the manufacturer's instructions.
|
| Label |
Cy3
|
| Label protocol |
Sample labeling was performed according to Agilent One-Color Microarray-Based Gene Expression Analysis protocol (Agilent Technology) with minor modifications. Briefly, mRNA was purified from total RNA after removal of rRNA (mRNA-ONLY™ Eukaryotic mRNA Isolation Kit, Epicentre). Then, each sample was amplified and transcribed into fluorescent cRNA along the entire length of the transcripts without 3' bias utilizing a mixture of oligo(dT) and random primers (Arraystar Flash RNA Labeling Kit, Arraystar). 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.
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| |
|
| Hybridization protocol |
One μg of each labeled cRNA was fragmented by adding 5 μl 10 × Blocking Agent and 1 μl of 25 × Fragmentation Buffer, then heated the mixture at 60°C for 30 min, finally 25 μl 2 × GE Hybridization buffer was added to dilute the labeled cRNA. 50 μl of hybridization solution was dispensed into the gasket slide and assembled to the LncRNA 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 |
PSC27 expression 7 days after treatment by 100 nM vincristine for 12 hrs
|
| Data processing |
Agilent Feature Extraction software (version 11.0.1.1) was used to analyze acquired array images. Quantile normalization,Combat between the groups of error correction and subsequent data processing were performed with using the GeneSpring GX v12.1 software package (Agilent Technologies).
After quantile normalization of the raw data, LncRNAs and mRNAs that at least 11 out of 21 samples have flags in Present or Marginal (“All Targets Value”) were chosen for further data analysis. Differentially expressed LncRNAs and mRNAs with statistical significance between the two groups were identified through P-value/FDR filtering. Differentially expressed LncRNAs and mRNAs between the two samples were identified through Fold Change filtering. Hierarchical Clustering and combined analysis were performed using homemade scripts.
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| |
|
| Submission date |
May 30, 2016 |
| Last update date |
Aug 31, 2018 |
| Contact name |
Yu Sun |
| E-mail(s) |
sunyu@sinh.ac.cn, submarinesuny@yahoo.com
|
| Phone |
86-21-54923302
|
| Organization name |
Chinese Academy of Sciences
|
| Department |
Shanghai Institute of Nutrition and Health
|
| Lab |
Cancer Resistance
|
| Street address |
320 Yueyang Rd, Life Sciences Building A #1508
|
| City |
Shanghai |
| ZIP/Postal code |
200031 |
| Country |
China |
| |
|
| Platform ID |
GPL16956 |
| Series (1) |
| GSE82033 |
Genome-wide expression profiling of human prostate stromal cells in response to multiple chemotherapeutic agents |
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