|
| Status |
Public on Feb 01, 2022 |
| Title |
Follicuar_exosome_Con |
| Sample type |
RNA |
| |
|
| Source name |
exosomes
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: ovarian follicular fluid
rna fraction: exosomal miRNA
disease state: non-polycystic ovary syndrome
|
| Treatment protocol |
Follicular fluid and blood samples were collected on the day of oocyte retrieval
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Exosomes were isolated using Exoquick kit purchased from System Biosciences following the manufacturer’s protocol. Exosomal miRNAs were isolated using the miRNeasy mini Kit purchased from Qiagen.
|
| Label |
cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 0.2 μg RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
0.6 μg of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/μg cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 22.5μl containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers' instructions. On completion of the fragmentation reaction, 22.5μl of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Human miRNA Expression（8*60K）for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505C) using one color scan setting for 4x180k array slides (Scan Area 61x21.6 mm, Scan resolution 3um, Dye channel is set to Green and Green PMT is set to 100%).
|
| Description |
miRNA
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.7.3 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities as the raw data. Raw data were normalized in quantile algorithm with Genespring 13.0(Agilent). Probe that at least 1 out of 2 samples flagged as Detected were maintained.
|
| |
|
| Submission date |
Feb 14, 2020 |
| Last update date |
Feb 01, 2022 |
| Contact name |
Chenchen Cui |
| E-mail(s) |
[email protected]
|
| Organization name |
Henan Provincial People’s Hospital
|
| Department |
Department of Reproductive Medicine Center
|
| Street address |
Weiwu Road #7
|
| City |
Zhengzhou |
| State/province |
Henan |
| ZIP/Postal code |
450003 |
| Country |
China |
| |
|
| Platform ID |
GPL21576 |
| Series (1) |
| GSE145318 |
Identification of exosomal miRNAs specifically derived from intrafollicular cells involved in polycystic ovary syndrome |
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