|
| Status |
Public on Jan 05, 2018 |
| Title |
ExVivoBlood_BPD_rep1_miRNA |
| Sample type |
RNA |
| |
|
| Source name |
Peripheral Blood, BPD, replicate 1
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: whole blood
gestational age: 28 weeks
diagnosis: Bronchopulmonary dysplasia (BPD)
|
| Treatment protocol |
A total of 20 premature infants with BPD according to the National Institute of Child Health and Human Development (NICHD) guidelines and 20 non-BPD age-matched controls were enrolled from clinic at department of neonatology in Shanghai Children Hospital.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using TRIzol (Invitrogen) and purified with RNeasy mini kit (QIAGEN) according to manufacturer’s instructions. RNA quality and quantity was measured by using nanodrop spectrophotometer (ND-1000, Nanodrop Technologies) and RNA Integrity was determined by gel electrophoresis.
|
| Label |
HY3
|
| Label protocol |
After quality control, the miRCURY™ Hy3™/Hy5™ Power labeling kit (Exiqon, Vedbaek, Denmark) was used according to the manufacturer’s guideline for miRNA labelling by following steps: a, 1μLRNA in 2.0 μL of water was combined with 1.0 μL of CIP buffer and CIP (Exiqon). The mixture was incubated for 30 min at 37°C. b, The Reaction was terminated by incubation for 5 min at 95°C. 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°C c, Terminated by incubation for 15 min at 65°C.
|
| |
|
| Hybridization protocol |
After stopping the labeling procedure, the Hy3™-labeled samples were hybridized on the miRCURYTM LNA Array (v.19.0) (Exiqon) according to array manual. a, The total 25 μL mixture from Hy3™-labeled samples with 25 μL hybridization buffer were first denatured for 2 min at 95°C, incubated on ice for 2 min.b, Then hybridized to the microarray for 16–20 h at 56°C in a 12-Bay Hybridization Systems (Hybridization System - Nimblegen Systems, Inc., Madison, WI, USA), Following hybridization, the slides were achieved, washed several times using Wash buffer kit (Exiqon)
|
| Scan protocol |
Then the slides were scanned using the Axon GenePix 4000B microarray scanner (Axon Instruments, Foster City, CA)
|
| Description |
B1
Gene expression in BPD human blood
|
| Data processing |
Scanned images were then imported into GenePix Pro 6.0 software (Axon) for grid alignment and data extraction. Replicated miRNAs were averaged and miRNAs that intensities>=30 in all samples were chosen for calculating normalization factor. Expressed data were normalized using the Median normalization. After normalization, significant differentially expressed miRNAs between two groups were identified through Fold change and P-value. Differentially expressed miRNAs between two samples were filtered through Fold change. Finally, hierarchical clustering was performed to show distinguishable miRNA expression profiling among samples.
The data matrix contains normalized signal intensity and is available on the series record.
|
| |
|
| Submission date |
Jan 04, 2018 |
| Last update date |
Jan 23, 2018 |
| Contact name |
cai cheng |
| E-mail(s) |
caicheng2004@163.com
|
| Organization name |
Shanghai Children’s Hospital, Shanghai Jiao Tong University
|
| Department |
Department of neonatology
|
| Street address |
No. 355, Luding Road
|
| City |
Shanghai |
| ZIP/Postal code |
200062 |
| Country |
China |
| |
|
| Platform ID |
GPL17107 |
| Series (2) |
| GSE108755 |
Differentially expressed genes in blood of BPD and Normal preterm neonates [miRNA] |
| GSE108756 |
Differentially expressed genes in blood of BPD and Normal preterm neonates |
|
