|
| Status |
Public on Jun 18, 2008 |
| Title |
E11.5_homozygous_embryo#7 |
| Sample type |
RNA |
| |
|
| Source name |
Ott1 knockdown, homozygous embryo #7
|
| Organism |
Mus musculus |
| Characteristics |
The mouse ES cell line XK135 (obtained from BayGenomics; funded by the National Heart, Lung, and Blood Institute, http://baygenomics.ucsf.edu), harbours a LacZ-neo fusion cassette inserted into the first coding exon of Ott1, 20 nt upstream the initiator codon, generating a low-expression Ott1XK135 allele. After injection into C57BL/6 blastocysts, the chimeric mice were bred for germline transmission. Genomic DNA from tail snips or yolk sacs was used for PCR genotyping. Ott1 alleles were detected by using primers, which span the XK135 insertion site.
|
| Treatment protocol |
Dissected embryos, snap-frozen in liquid nitrogen
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted and purified using RiboPure RNA isolation (Ambion).
Total RNA samples were quantitated by UV spectrophotometry (OD260/280).
Quality of Total RNA was assessed using an Agilent Bioanalyzer.
|
| Label |
biotin
|
| Label protocol |
First and second strand cDNA was prepared from the total RNA samples using standard CodeLink protocol.
Biotinylated cRNA target was prepared from the DNA template and verified on the Bioanalyzer.
cRNA was fragmented to uniform size and verified on the Bioanalyzer.
|
| |
|
| Hybridization protocol |
CodeLink Bioarrays were hybridized with the cRNA target and stained with Cy5-streptavidin.
|
| Scan protocol |
Slides were washed and then scanned on an Axon GenePix 4000B scanner.
|
| Description |
Mice harboring a low expression Ott1 allele were mated and embryos at day E11.5 were dissected and genotyped.
|
| Data processing |
Data were analyzed for quality with CodeLink software package, and transcripts of which the values were missing in at least one mictoarray were removed from the dataset, and raw intensity values less than 0.01 were set to 0.01.
Normalization Method: Median-Normalization;
Computation Method for Median: Spot Based;
Threshold Trim Percentage: 20;
Raw TrimMean Negative Control: 16.26547943;
Normalized TrimMean Negative Control: 0.26178759;
Median: 62.13235474
|
| |
|
| Submission date |
Jun 13, 2008 |
| Last update date |
Jun 17, 2008 |
| Contact name |
Andrei S Zolotukhin |
| E-mail(s) |
zolotukh@ncifcrf.gov
|
| Organization name |
NCI-Frederick
|
| Street address |
P.O. Box B
|
| City |
Frederick |
| State/province |
MD |
| ZIP/Postal code |
21702 |
| Country |
USA |
| |
|
| Platform ID |
GPL2897 |
| Series (1) |
| GSE11785 |
Control of gene expression by Ott1/RBM15 protein |
|
