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| Status |
Public on Jun 05, 2021 |
| Title |
BxPC3_dau_72h_3 [miRNA] |
| Sample type |
RNA |
| |
|
| Source name |
BxPC3_dau_72h
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: BxPC3
cell type: Human pancreatic adenocarcinoma cells
treatment: 24 μmol Dau for 72h
tag: Human pancreatic adenocarcinoma cells
|
| Treatment protocol |
After the cells grew to 80% of the culture dish, add 24μmol dau for 72h.
|
| Growth protocol |
BxPC3 cells were cultured in RPMI-1640 (Hyclone, USA) supplemented with 10% fetal bovine serum (FBS) and 1% penicillin streptomycin solution in an atmosphere of 95% air and 5% CO2 in a 37℃ incubator.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol extraction of total RNA was performed according to the manufacturer's instructions.
|
| Label |
Hy3
|
| Label protocol |
Briefly, total RNA from each sample was labeled with Hy3 according to miRCURYTM Array Power Labeling kit (Cat #208032-A, Exiqon) manufactor
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| |
|
| Hybridization protocol |
1. Kits and Reagents: 2×Hybridization buffer; Phalanx Hyb. Assembly miRCURYTM Array, Wash buffer kit (Cat #208021, Exiqon):Buffer A, Buffer B,Buffer C 2. Prepare the hybridization mix in a PCR tube 3. Incubate at 95°C for 2 min. During the incubation the target preparation should be protected from light. 4. Leave on ice for at least 2 min. and up to 15min. Briefly spin the reaction after ice incubation. 5. Take cover slide out of the package. 6. Carefully laid the miRCURYTM Array on top of the assembly of cover slide and spacer, where the printed side of array should be facing toward the cover slide, to form a hybridization assembly. The printed side of array is on the side of the label. 7. Insert the hybridization assembly into a (3.1 x 9cm) heat-shrank hybridization (hyb.) bag. Sample loading side of the hybridization assembly should face the opening side of film bag. Lower the assembly to the end of the bag (Assembly viewed from the other side). 8. Clip the bag with a clipper securely from the opening end. Immerse the assembly to the rim of the slide in 95℃ hot water swiftly (approximately 2~5 sec). Do not dip the assembly too far into the water to avoid water leaking into the assembly. The hyb. bag will shrink and tightly wrap around the assembly. 9. Remove the assembly from water and wipe off the water from the assembly. Trim off the excess film from the top of the assembly. Keep the assembly in a 50℃ oven for at least 10 minutes. 10. Load the180ul target hybridization mix into the assembly through the sample loading opening. Fill up the hybridization space with 1X Hybridization buffer till the liquid level reaches to approximately 5 mm (0.5cm) away from the top rim. 11. Cut the second Hyb. Bag to approximately one-half of its original length. 12. Keep the assembly vertical and slip on the shortened hyb. bag from step 11 all the way until the assembly reaches the end of the hyb. bag. 13. Use a pair of forceps to hold onto the top of the assembly. Immerse the entire assembly into 95℃ hot water with the assembly in upright position. 14. Place the hybridization assembly into a 56℃ oven and set it on a rotator. Remember to rotate the assembly at 2 rotations per minute (RPM) for overnight. 15. After hybridization, disassembled the hybridization assembly, and wash the slides at 56℃ for 2min. using Wash buffer A. 16. Wash briefly at room temperature in Wash buffer B. 17. Wash for 2min. at room temperature in Wash buffer B. 18. Wash for 2min. at room temperature in Wash buffer C. 19. Wash briefly in water. 20. Dry the slides by centrifugation for 5 min. at 200* g(1000rpm). Scan slides immediately.
|
| Scan protocol |
Scanning is performed with the Axon GenePix 4000B microarray scanner. GenePix pro V6.0 is used to read the raw intensity of the image.
|
| Description |
After the cells grew to 80% of the culture dish, add 24μmol dau for 72h.
|
| Data processing |
a) The intensity of green signal is calculated after background subtraction and four Replicated spots of each probe on the same slide have been averaged. b) We use Median Normalization Method to obtain Normalized Data, Normalized Data= (Foreground-Background)¬/¬median, the median is 50 percent quantile of microRNA intensity which is larger than 30 in all samples after background correction. c) After normalization, the statistical significance of differentially expressed miRNA was analyzed by T-test. d) Unsupervised hierarchical clustering and Correlation analysis was performed on miRNA data. e) You can find the detail information in miRNA Expression Profiling Data.
nomalized_data.txt contains Normalized signal intensity data
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| |
|
| Submission date |
Jun 04, 2021 |
| Last update date |
Jun 05, 2021 |
| Contact name |
Yun Bai |
| E-mail(s) |
[email protected]
|
| Phone |
+8613304642998
|
| Organization name |
Hei Long Jiang University of Chinese medicine
|
| Street address |
No.24 heping Street, XiangFang district
|
| City |
Harbin |
| State/province |
Hei Long Jiang |
| ZIP/Postal code |
150040 |
| Country |
China |
| |
|
| Platform ID |
GPL17728 |
| Series (2) |
| GSE176163 |
The study of the effects of dauricine(dau) on human pancreatic carcinoma cells [miRNA] |
| GSE176164 |
The study of the effects of dauricine(dau) on human pancreatic carcinoma cells |
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