|
| Status |
Public on Apr 26, 2022 |
| Title |
non-heparanized blood, Ct-postive, spontaneous abortion, rep1 |
| Sample type |
RNA |
| |
|
| Source name |
non-heparanized blood
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: serum
gender: female
age: 26y
infection: Ct-postive
history: recurrent spontaneous aborter (RSA)
|
| Treatment protocol |
Freshly drawn blood from recurrent spontaneous aborters and controls were collected. Serum was separated and stored for further use.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using Trizol(Ambion). RNA was quantified using a NanoDrop-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
|
| Label |
Cy3
|
| Label protocol |
200ng of total RNA was dephosphorylated using Calf Intestinal Alkaline Phosphatase (CIP) master mix
The dephosphorylated miRNA sample was denatured by adding Dimethyl Sulfoxide
and heating at 100 oC for 10 minutes and transferred to ice-water bath. The Ligation master mix (Agilent Technologies, Part Number: 5190-0456) containing Cyanine 3-pCp was added to the denatured miRNA sample and incubated at 16oC for 2 hours. The Cyanine 3-pCp labeled miRNA sample was dried completely in the vacuum concentrator (Eppendorf, Concentrator Plus, Catalog Number 5305000) at 45oC for 2 hour
|
| |
|
| Hybridization protocol |
The dried sample was resuspended in nuclease free water and mixed with Hybridization Mix containing blocking solution (Agilent Technologies, Part Number: 5190-0456) and Hi-RPM Hybridization Buffer (Agilent Technologies, Part Number: 5190-0456) and incubated at 100oC for 5 minutes followed by snap chill on ice for 5 minutes. The samples were hybridized on the Human miRNA 8x60K Arrays. The hybridization was carried out at 55°C for 20 hours.
After hybridization, the slides were washed using Gene Expression Wash Buffer1 (Agilent Technologies, Part Number 5188-5325) at room temperature for 5 minutes and Gene Expression Wash Buffer 2 (Agilent Technologies, Part Number 5188-5326) at 37oC for 5 minutes.
|
| Scan protocol |
The microarray slide was scanned on a G2600D scanner (Agilent Technologies). Data extraction from Images was done using Feature Extraction software v 11.5.1.1 of Agilent.
|
| Description |
miRNA expression
|
| Data processing |
Data extraction from Images was done using Feature Extraction software v 11.5.1.1 of Agilent.
|
| |
|
| Submission date |
Apr 25, 2022 |
| Last update date |
Apr 26, 2022 |
| Contact name |
Ankita Ray |
| E-mail(s) |
[email protected]
|
| Phone |
9717743274
|
| Organization name |
ICMR-National Institute of Pathology
|
| Lab |
Molecular Microbiolgy laboratory
|
| Street address |
Safdarjung hospital campus
|
| City |
New Delhi |
| State/province |
Delhi |
| ZIP/Postal code |
110029 |
| Country |
India |
| |
|
| Platform ID |
GPL21576 |
| Series (1) |
| GSE201442 |
Microarray profiling of microRNA in Chlamydia trachomatis-infected recurrent spontaneous aborters |
|
