|
| Status |
Public on Apr 10, 2020 |
| Title |
Patient 3 A3_hlm |
| Sample type |
RNA |
| |
|
| Source name |
normal liver samples
|
| Organism |
Homo sapiens |
| Characteristics |
Sex: male
age: 58 years
tissue source: normal liver
tissue: bone marrow
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA were extracted using Trizol. The 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 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. 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. The hybridized arrays were washed, fixed and scanned using the Agilent Scanner G2505C
|
| |
|
| 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 × 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)
|
| Data processing |
Data was extracted using Agilent Feature Extraction software.A series of data processing including quantile normalization were performed using the R software limma package. The circRNAs that at least 3 out of 6 samples have flags in P or M (defined by GeneSpring software) were retained for further differential analyses.Differentially expressed circRNAs with statistical significance between two samples or two groups were identified using fold change cutoff or through Volcano Plot filtering respectively.The circRNA/microRNA interaction was predicted with Arraystar's home-made miRNA target prediction software. All the differentially expressed circRNAs were annotated in detail with the circRNA/miRNA interaction information. .
|
| |
|
| Submission date |
Jun 20, 2019 |
| Last update date |
Apr 10, 2020 |
| Contact name |
FU YUN FENG |
| E-mail(s) |
14579@qq.com
|
| Phone |
13786450644
|
| Organization name |
The Third Xiangya Hospital of Central South University
|
| Department |
Blood
|
| Lab |
Blood
|
| Street address |
No. 138 Tonglu Road, Yuelu District, Hexi Distric
|
| City |
CHANG SHA |
| State/province |
HUNAN |
| ZIP/Postal code |
410000 |
| Country |
China |
| |
|
| Platform ID |
GPL21825 |
| Series (1) |
| GSE133058 |
hsa_circ_0007841: A novel potential biomarker and drug resistance for multiple myeloma |
|
