|
| Status |
Public on Aug 27, 2020 |
| Title |
L-PCa4 |
| Sample type |
RNA |
| |
|
| Source name |
prostate cancer Gleason<6
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: prostate cancer
gender: male
|
| Treatment protocol |
Place a freshly dissected cross-section prostate tissue (5mm thick) on to a pre-labeled tissue base mold. Cover the entire tissue block with Surgipath FSC 22 Frozen Section Embedding Medium (Leica). Slowly place the base mold containing the tissue block into liquid nitrogen till the entire tissue block is submerged into liquid nitrogen to ensure tissue is frozen completely. Store the frozen tissue block at -80°C until ready for sectioning. Transfer the frozen tissue block to a cryotome cryostat (-20 °C) prior to sectioning and allow the temperature of the frozen tissue block to equilibrate to the temperature of the cryotome cryostat. Section the frozen tissue block into 8 μm using the cryotome.Place the tissue sections onto glass slides suitable for HE staining and immunohistochemistry.The tissue specimens were confirmed by two experienced pathologists. Pathological grading was judged by Gleason points-scoring system, Gleason score ≥8 and Gleason score <8.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA extraction was performed with the miRNeasy Mini Kit (217004; Qiagen) according to the manufacturer's instructions. The purity and concentration of total RNA samples were determined with NanoDrop ND-1000. Results were provided in Sample QC report.
|
| Label |
Cy3
|
| Label protocol |
Sample labeling and array hybridization were performed according to the manufacturer’s protocol (Arraystar Inc.). Briefly, total RNAs were digested with Rnase R (Epicentre, Inc.) to remove linear RNAs and enrich circular RNAs. Then, the enriched circular RNAs were amplified and transcribed into fluorescent cRNA utilizing a random priming method (Arraystar Super 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.
|
| |
|
| Hybridization protocol |
1 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 × Hybridization buffer was added to dilute the labeled cRNA. 50 μl of hybridization solution was dispensed into the gasket slide and assembled to the circRNA expression microarray slide. The slides were incubated for 17 hours at 65°C in an Agilent Hybridization Oven.
|
| Scan protocol |
After washing, slides were scanned with the Agilent Scanner G2505C. Scanned images were imported into Agilent Feature Extraction software for raw data extraction.
|
| Description |
circular RNA
|
| Data processing |
Agilent Feature Extraction software (version 11.0.1.1) was used to analyze acquired array images. Quantile normalization and subsequent data processing were performed using the R software limma package. Differentially expressed circRNAs with statistical significance between two groups were identified through Volcano Plot filtering. Differentially expressed circRNAs between two samples were identified through Fold Change filtering. Hierarchical Clustering was performed to show the distinguishable circRNAs expression pattern among samples.
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| |
|
| Submission date |
Apr 16, 2018 |
| Last update date |
Aug 27, 2020 |
| Contact name |
Zhan Yang |
| E-mail(s) |
benky2010@qq.com
|
| Phone |
+8631166002811
|
| Organization name |
The Second Hospital of Hebei Medical University
|
| Street address |
215 Heping W Rd
|
| City |
Shijiazhuang |
| State/province |
Hebei |
| ZIP/Postal code |
050000 |
| Country |
China |
| |
|
| Platform ID |
GPL21825 |
| Series (2) |
| GSE113153 |
Dysregulation of p53-RBM25-mediated circAMOTL1L biogenesis contributes to prostate cancer progression (Agilent circRNA data set) |
| GSE113180 |
Dysregulation of p53-RBM25-mediated circAMOTL1L biogenesis contributes to prostate cancer progression |
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