|
| Status |
Public on Oct 01, 2014 |
| Title |
Male CSF2 vs Control Rep. 1 |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
Day 15 EEM from Male Control Rep. 1
|
| Organism |
Bos taurus |
| Characteristics |
embryo: Filamentous
tissue: whole embryo
treatment: control
|
| Treatment protocol |
10 ng/ml of CSF2 or Control (DPBS/BSA) from Day 5-7 post-insemination during in vitro culture of bovine embryos.
|
| Growth protocol |
Fertilized embryos were grown in culture to Day 7 after fertilization and transferred into Holstein recipients. At Day 15, they were flushed from the recipient using DPBS/PVP
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
DNA was extracted according to manufacturer's instructions using the Qiagen Allprep DNA/RNA mini kit
|
| Label |
Cy3
|
| Label protocol |
DNA was labeled using the Universal Linkage System (ULS) by Kreatech for gene expression analysis. Microarray slides were made by Agilent and designed by EmbryoGene. ULS directly labels the aRNA through a non-enzymatic protocol which labels through the formation of a coordinate bond on the N7 position of guanine. The Cy3 and Cy5 are linked by a spacer to the ULS molecule.
|
| |
|
| Channel 2 |
| Source name |
Day 15 EEM from Male CSF2 Rep. 1
|
| Organism |
Bos taurus |
| Characteristics |
tissue: whole embryo
embryo: Filamentous
treatment: CSF2
|
| Treatment protocol |
10 ng/ml of CSF2 or Control (DPBS/BSA) from Day 5-7 post-insemination during in vitro culture of bovine embryos.
|
| Growth protocol |
Fertilized embryos were grown in culture to Day 7 after fertilization and transferred into Holstein recipients. At Day 15, they were flushed from the recipient using DPBS/PVP
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
DNA was extracted according to manufacturer's instructions using the Qiagen Allprep DNA/RNA mini kit
|
| Label |
Cy5
|
| Label protocol |
DNA was labeled using the Universal Linkage System (ULS) by Kreatech for gene expression analysis. Microarray slides were made by Agilent and designed by EmbryoGene. ULS directly labels the aRNA through a non-enzymatic protocol which labels through the formation of a coordinate bond on the N7 position of guanine. The Cy3 and Cy5 are linked by a spacer to the ULS molecule.
|
| |
|
| |
| Hybridization protocol |
one Cy3 labeled DNA sample and one Cy5 DNA sample were combined and hybridized to one area of a 2x444,000 Agilent EmbryoGene microarray slide. 635=Cy5 and 532=Cy3.
|
| Scan protocol |
Slides were scanned using a Agilent High-resolution C Scanner (Agilent Technologies, CA, USA) while features were extracted using Agilent's feature extraction software (Agilent)
|
| Data processing |
Raw data were normalized using within-array loess normalization followed by between-array quantile normalization. Normalized data were then fit to a linear model before being subjected to an empirical Bayes correction. Mean intensity of each spot on the array was transformed by log2 while p-values were determined using a moderated t-statistic (60). Regions with an estimated absolute fold-change above 1.5 or below 0.67 and a p-value < 0.05 were considered as differentially methylated.
|
| |
|
| Submission date |
Mar 13, 2014 |
| Last update date |
Oct 01, 2014 |
| Contact name |
Kyle Dobbs |
| E-mail(s) |
[email protected]
|
| Organization name |
University of Florida
|
| Department |
Animal Sciences
|
| Street address |
2250 SW Shealy Drive
|
| City |
Gainesville |
| State/province |
Florida |
| ZIP/Postal code |
32611 |
| Country |
USA |
| |
|
| Platform ID |
GPL18384 |
| Series (1) |
| GSE55873 |
Bovine Day 15 Extra-embryonic membrane (EEM): Male and Female Control vs. Male and Female CSF2 Methylation |
|
