|
| Status |
Public on Mar 14, 2018 |
| Title |
Serum_DOR_rep2 |
| Sample type |
RNA |
| |
|
| Source name |
DOR serum
|
| Organism |
Homo sapiens |
| Characteristics |
disease state: DOR
sample type: serum
age: 34 years
afc: 3
fsh: 14.13
lh: 5.23
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using TRIzol LS (Invitrogen) and purified with RNeasy mini kit (QIAGEN) according to manufacturer’s instructions. RNA quality and quantity was measured by using nanodrop 1000. RNA integrity was determined by gel electrophoresis.
|
| Label |
Hy3
|
| Label protocol |
The miRCURYTM Hy3TM/ Hy5TM Power labeling kit (Exiqon) was used according to the manufacturer’s guideline for miRNA labelling by following steps: 1 µl RNA in 3.0 µl of water was combined with 1.0 µl of CIP buffer and CIP (Exiqon). The mixture was incubated for 30 min at 37℃. The Reaction was terminated by incubation for 5 min at 95℃. Then 3.0 µl of labeling buffer, 1.5 µl of fluorescent label (Hy3TM), 2.0 µl of DMSO, 2.0 µl of labeling enzyme were added into the mixture. The labeling reaction was incubated for 1 h at 16℃. Terminated by incubation for 15 min at 65℃.
|
| |
|
| Hybridization protocol |
The Hy3TM-labeling samples were hybridized on the miRCUYTM LNA Array (v,16.0) (Exiqon) according to array annual. The total 12.5 µl Hy3TM-labeled samples, 90 µl 2 × Hybridization buffer, 77.5 µl Nuclease-free Buffer were first denatured for 2 min at 95℃, incubated on ice for 2 min. Then hybridized to the microarray for 16-20 h at 56℃. Following hybridization, the slides were achieved, washed several times Wash buffer kit (Exiqon).
|
| 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 |
The 6th generation of miRNA array contains more than 1891 capture probes, covering all human, mouse and rat microRNAs annotated in miRBase 16.0, as well as all viral microRNAs related to these species. In addition, this array contains capture probes for 66 new miRPlus™ human microRNAs.
|
| Data processing |
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. So, if "Foreground-Background" is a negative number, the "Normalized Data" was determined to be "null". After normalization, the statistical significance of differentially expressed miRNA was analyzed by T-test.
GenePix Pro 6.0 software (Axon) was used for grid alignment and data extraction.
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| |
|
| Submission date |
Mar 13, 2018 |
| Last update date |
Mar 14, 2018 |
| Contact name |
XiaoKui Yang |
| Organization name |
Beijing Obstetrics and Gynecology Hospital, Capital Medical University
|
| Street address |
251, Yaojiayuan Road, Chaoyang DIstrct
|
| City |
Beijing |
| ZIP/Postal code |
100026 |
| Country |
China |
| |
|
| Platform ID |
GPL11434 |
| Series (2) |
| GSE111773 |
Differentially expressed miRNAs in serum of diminished ovarian reserve (DOR) patients |
| GSE111774 |
MiR-106a increases granulosa cell viability and is down-regulated in women with diminished ovarian reserve |
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