|
| Status |
Public on Oct 11, 2011 |
| Title |
C33A_24h_10Amol/L5AZA |
| Sample type |
RNA |
| |
|
| Source name |
10Amol/L5AZA_treated
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: C33A
cell type: cervical cancer cells
|
| Treatment protocol |
Four cervical cancer cells were choosed for this study,No.1 samples were treated with 5-azacytidine for 24hrs,the other samples were untreated , then cultured with RPMI 1640 medium supplemented with 10% heat-inactivated fetal bovine serum for 48 hours at 37 ℃ in a humidified incubator with 5% CO2 .
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was prepared using the trizol® Reagent (Invitrogen life technologies) following the manufacturer's recommendations. The protocol includes differential lysis of C33A, CASKI, HeLa, and SIHA cells, and RNasey Mini Kit(Qiagen p/n 74104). RNA quantification and quality control used a NanoDrop-1000 spectrophotometer.
|
| Label |
Hy3
|
| Label protocol |
Hy3™ labeled cRNA was prepared from 1 ug RNA using the miRCURYTM Array Power Labeling kit (Cat #208032-A, Exiqon) according to the manufacturer's instructions. Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
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| |
|
| Hybridization protocol |
12.5 ul of Hy3-labelled sample mixed with 2×Hybridization buffer and Nuclease-free Buffer in a reaction volume of 180 ul was Prepare the hybridization mix in a PCR tube.Load the 180ul target hybridization mix into the hybridization assembly through the sample loading opening .Place the hybridization assembly into a 56C oven and set it on a rotator at 2 rotations per minute (RPM) for overnight.After hybridization, disassembled the hybridization assembly, and wash the slides at 56C for 2min. using Wash buffer A.Wash for 2min. at room temperature in Wash buffer B and C.Wash briefly in water.Dry the slides by centrifugation for 5 min at 200g (1000rpm).
|
| Scan protocol |
Slides were scanned immediately after washing on the Axon GenePix 4000B microarray scanner.
|
| Description |
Gene expression after 24hr in 10 Amol/L 5-aza-2'-deoxycytidine
|
| Data processing |
GenePix pro V6.0 is used to read the raw intensity of the image.The intensity of green signal is calculated after background subtraction and four Replicated spots of each probe on the same slide have been averaged. Median Normalization Method was used to obtain 'Normalized Data', Normalized Data = (Foreground-Background)/median, the median is 50 percent quantile of microRNA intensity which is larger than 50 in all samples after background correction. After normalization, the statistical significance of differentially expressed miRNA was analyzed by T-test. Unsupervised hierarchical clustering and Correlation analysis was performed on miRNA data.
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| |
|
| Submission date |
Oct 10, 2011 |
| Last update date |
Oct 11, 2011 |
| Contact name |
yao ting ting |
| E-mail(s) |
squall41269@21cn.com
|
| Phone |
008613265991256
|
| Fax |
00862081332199
|
| Organization name |
SUN YAT-SEN MEMORIAL,HOSPITAL,SUN YAT-SEN UNIVERSITY
|
| Street address |
107 Riverside Road
|
| City |
guangzhou |
| ZIP/Postal code |
510000 |
| Country |
China |
| |
|
| Platform ID |
GPL7723 |
| Series (1) |
| GSE32724 |
MicroRNA signatures in human cervical cancer |
|
