|
| Status |
Public on Feb 26, 2019 |
| Title |
Disease free Endometrium of fertile women, replicate 2 |
| Sample type |
RNA |
| |
|
| Source name |
human endometrium, Fertile Secretory without Endometriosis (Group 1A)
|
| Organism |
Homo sapiens |
| Characteristics |
gender: Female
sample group: Disease free Endometrium of fertile women
tissue: Endometrium
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was prepared using Trizol reagent. The protocol included brief homogenisation using polytron homogeniser, followed by phase seperation by chloroform and precipitation using iso-propanol. This was followed by washing by 75% ethanol and elution in nuclease-free waster. Total RNA was quantified using a NanoDrop-2000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer.
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 200 ng total RNA using the One-Color Low Input TOTAL RNA Linear Amplification kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (Qiagen). Dye incorporation and cRNA yield were checked with the NanoDrop ND-2000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
1.65 ug of Cy3-labelled cRNA (specific activity >6 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 55 ul containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 55 ul of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to 4 x 44k Agilent Whole Human Genome Oligo Microarrays 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 (G2505B) using one color scan setting for 4x44k array slides (Scan Area 61x21.6 mm, Scan resolution 5 um, Dye channel was set to Green, and Green PMT was set to 100%).
|
| Description |
SAMPLE 2
Homo sapiens whole genome expression microarray for human endometrium
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.7.3.1 (Agilent) using default parameters (protocol GE1 and Design Id: 014850) to obtain background subtracted and spatially detrended Processed Signal intensities. Features were flagged in Feature Extraction and Non-uniform Features and outliers were excluded.
|
| |
|
| Submission date |
Sep 18, 2018 |
| Last update date |
Feb 26, 2019 |
| Contact name |
Muzaffer Ahmed Bhat |
| E-mail(s) |
muzaffar.sciml87@gmail.com
|
| Phone |
91 8826753204
|
| Organization name |
AIIMS
|
| Department |
Physiology
|
| Lab |
Molecular Biology
|
| Street address |
ANSARI NAGAR NEW DELHI 110029
|
| City |
Delhi |
| State/province |
Delhi |
| ZIP/Postal code |
110029 |
| Country |
India |
| |
|
| Platform ID |
GPL6480 |
| Series (1) |
| GSE120103 |
Homo sapiens whole genome expression microarray of endometrium obtained from fertile and Infertile women with stage IV ovarian endometriosis and without endometriosis |
|
