|
| Status |
Public on Feb 13, 2019 |
| Title |
iPax7 re-isolated microarray rep3 |
| Sample type |
RNA |
| |
|
| Source name |
iPax7 re-isolated
|
| Organism |
Mus musculus |
| Characteristics |
strain: NSG/NSGmdx4cv
tissue: Tibialis anterior
cell type: mononucleated cells
age: 3 months-old
|
| Extracted molecule |
total RNA |
| Extraction protocol |
1500 cells isolated from 3 mice/line in 3 independent experiments were sorted directly into the SuperAmp™ lysis buffer (Miltenyi Biotech)
|
| Label |
Cy3
|
| Label protocol |
RNA was amplified and retrotranscribed according to SuperAmp™ Preparation kit’s instruction. cDNA was quantified and analyzed on a Agilent 2100 Bioanalyzer platform (AgilentTechnologies). 250 ng of each cDNA were used as template for Cy3 labeling which was performed according to Miltenyi Biotec’s undisclosed protocol.
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| |
|
| Hybridization protocol |
The Cy3- labeled cDNAs were hybridized overnight (17 hours, 65°C) to an Agilent Whole Mouse Genome Oligo Microarrays 4 x 44K using Agilent’s recommended hybridization chamber and oven. Microarrays were then washed once with 6x SSPE buffer containing 0.005% N-lauroylsarcosine for 1 min at room temperature followed by a second wash with pre-heated 0.06x SSPE buffer (37 °C) containing 0.005% N-lauroylsarcosine for 1 min.
|
| Scan protocol |
Fluorescence signals of the hybridized Agilent Microarrays were detected using Agilent’s Microarray Scanner System (Agilent Technologies). Finally, the Agilent Feature Extraction Software (FES) was used to read out and process the microarray image files.
|
| Description |
iPax7_MNC_3
|
| Data processing |
Microarrays were analyzed in R (3.4.3) using the limma package. First, background correction was performed on each microarray using “normexp” method. Then we performed quantile normalization between all microarrays before performing differential expression analysis. Filtering on microarray was conducted by retaining only the probes that were above background in at least 4 arrays.
For differential expression analysis, we first fitted a linear model on the normalized microarray expression and then used an empirical Bayes moderation approach to calculate the statistical significance. The differentially expressed genes are defined as fold change over 2 and false-discovery-rate adjusted p-value less than 0.05. Hierarchical clusterings were performed using correlation as similarity metric and average-link method in R.
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| |
|
| Submission date |
Oct 22, 2018 |
| Last update date |
Feb 13, 2019 |
| Contact name |
Tania Incitti |
| E-mail(s) |
tincitti@umn.edu
|
| Organization name |
University of Minnesota
|
| Street address |
2231 6th Street SE
|
| City |
Minneapolis |
| State/province |
Minnesota |
| ZIP/Postal code |
55455 |
| Country |
USA |
| |
|
| Platform ID |
GPL4134 |
| Series (2) |
| GSE121615 |
Pluripotent stem cell-derived myogenic progenitors remodel their molecular signature upon in vivo engraftment [Microarray] |
| GSE121639 |
Pluripotent stem cell-derived myogenic progenitors remodel their molecular signature upon in vivo engraftment |
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