|
| Status |
Public on Nov 18, 2011 |
| Title |
wt ovary 1 |
| Sample type |
RNA |
| |
|
| Source name |
wild-type
|
| Organism |
Drosophila melanogaster |
| Characteristics |
genotype: wild-type
tissue: ovary
|
| Treatment protocol |
none
|
| Growth protocol |
Adult 2-4 day old female flies were fed yeast paste and maintained at 25ºC.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The RNA was purified using Qiagen RNAeasy kit following the manufacturer's protocol(Qiagen)
|
| Label |
biotin
|
| Label protocol |
Biotin
|
| |
|
| Hybridization protocol |
Recommended Affymetrix protocol
|
| Scan protocol |
The tiling arrays were scanned using an Affymetrix GeneChip Scanner 3000 7G according to manufacturer's instruction.
|
| Data processing |
Since transposons account for more than 10% of the probes on the tiling array, a special normalization workflow has been taken to avoid over-normalization of signals. Coordinates and raw signal values for Drosophila tiling 2.0R array probes were extracted from Affymetrix Tiling Array Software. Then probes mapped to transposon regions were identified. The remaining non-transposon probes were quantile-normalized across qin mutant replicates based on the assumption that the overall signal distribution of these probes should remain the same across the different strains. Then the signals for transposon probes were calculated by looking up the normalized values of non-transposon probes at same raw signal level. To summarize the signal for each transposon element, probes mapped to any copy of the element were grouped together and Hodges-Lehmann estimator (ref1) was used to calculate the pseudomedian of their normalized signals. We used pseudomedian as it is less sensitive to the large number of outlier probes (probes with value of 1) in the tiling array experiments. Differentially expressed transposon elements were identified by contrasting their pseudomeidan values in three mutant replicates against three wild type replicates (Klattenhoff et al., 2009, Cell, 138,1137-49. ( accession number: GSE14307)). To correct for multiple testing, Express changed more than 3–fold and False Discovery Rates (FDRs) were calculated from t-test P-values. FDR<0.1 were used to call significantly changed transposons. Transposons with average expression value less than a minimal expression threshold of 149 (the 50th percentile expression value of RefSeq mRNAs) in both wild type and mutants were flagged as they are at the detection limit of the tiling array. Expression values of mRNAs were summarized by calculating the pseudomedian of probes mapped to each of the RefSeq mRNA transcripts. Differentially expressed transcripts were identified using FDR<0.1 cutoff.
|
| |
|
| Submission date |
Jun 17, 2011 |
| Last update date |
Nov 18, 2011 |
| Contact name |
Zhao Zhang |
| E-mail(s) |
zhao.zhang@umassmed.edu
|
| Phone |
508-918-9608
|
| Organization name |
UMass Medical School
|
| Department |
BMP
|
| Lab |
Zamore/Theurkauf
|
| Street address |
LRB-870N, 364 Plantation Street
|
| City |
Worcester |
| State/province |
MA |
| ZIP/Postal code |
01605 |
| Country |
USA |
| |
|
| Platform ID |
GPL6629 |
| Series (1) |
| GSE30061 |
Characterization of expression changes in qin by tiling array |
|
