|
| Status |
Public on Nov 13, 2018 |
| Title |
w;tim-GAL4(UAS)/UAS-miR-210, biological replicate 2 (miR-210-ZT0_over_2) |
| Sample type |
RNA |
| |
|
| Source name |
miR-210 overexpressing_adult flies_brain
|
| Organism |
Drosophila melanogaster |
| Characteristics |
genotype/variation: w;tim-GAL4(UAS)/UAS-miR-210
developmental stage: adult flies
time point: ZT0
tissue: brain
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from approximately 25 flies brains for each genotype using ZR RNA MicroPrepTM (ZYMO RESEARCH) according to the manufacter's instructions. All RNA samples were checked for quality by capillary electrophoresis (RNA 6000 Nano LabChip, Agilent Bioanalyzer 2100, Agilent Technologies).
|
| Label |
Cy3
|
| Label protocol |
50 ng of total RNA were labeled with “Low Input Quick Amp Labeling Kit, one color” according to the manufacturer’s instructions. The synthesized cDNA was transcribed into cRNA and labeled with Cy3-dCTP. Labeled cRNA was purified with RNeasy Mini columns (Qiagen, Valecia, CA). The quality of each cRNA sample was verified by total yield and specificity calculated with NanoDrop ND-1000 spectrophotometer measurements (Nanodrop, Wilmington, DE).
|
| |
|
| Hybridization protocol |
600 ng of labeled cRNA were used in each reaction and the hybridization was carried out at 65°C for 17 hours in an hybridization oven rotator (Agilent Technologies, Palo Alto, CA). The arrays were washed by Agilent Gene expression washing buffers and Stabilization and Drying Solution as suggest by the supplier.
|
| Scan protocol |
Slides were scanned on an Agilent microarray scanner (model G2565CA) and Agilent Feature Extraction software version 10.5.1.1 was used for image analysis.
|
| Description |
Gene expression of miR-210 overexpressing flies (miR-210-ZT0_over_2).
|
| Data processing |
Inter-array normalization of expression levels was performed with quantile method to correct for possible experimental distortions. A normalization function was applied to the expression data of all the experiments and the values of within-arrays replicate spots were then averaged. Feature Extraction Software, which provided spot quality measures, was used to evaluate the quality and reliability of the hybridization. In particular, the flag glsPosAndSignif (set to 1 if the spot had an intensity value significantly different from the local background and to 0 when otherwise) was used to filter out unreliable probes: the flag equal to 0 was noted as null. Probes with a high proportion of null values were removed from the dataset in order to carry out a more solid, unbiased, statistical analysis. Fifty percent of null was used as the threshold in the filtering process, and a total of 23,717 Drosophila transcripts were obtained.
|
| |
|
| Submission date |
Dec 17, 2015 |
| Last update date |
Nov 13, 2018 |
| Contact name |
Cristiano De Pitta |
| E-mail(s) |
[email protected]
|
| Phone |
+390498276210
|
| Organization name |
University of Padova
|
| Department |
Biology
|
| Street address |
Via U. Bassi, 58/B
|
| City |
Padova |
| ZIP/Postal code |
35131 |
| Country |
Italy |
| |
|
| Platform ID |
GPL18767 |
| Series (2) |
| GSE76099 |
Gene expression analysis of miR-210 over-expressing flies at ZT0. |
| GSE77245 |
Gene expression analysis of miR-210 over-expressing flies at ZT0 and ZT12 |
|
