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| Status |
Public on Feb 01, 2011 |
| Title |
Empirical Annotation of the Daphnia pulex genome; Experiment B |
| Organism |
Daphnia pulex |
| Experiment type |
Expression profiling by genome tiling array
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| Summary |
Experiments conducted on this tiling array are used to (1) validate the frozen gene sets of the current genome annotation, (2) improve the predicted gene structures by empirically determining UTRs and intron-exon boundaries, identifying missing upstream, internal, and downstream exons and alternative transcripts, (3) propose gene structure models in transcribed regions containing no predicted genes and (4) delineate transcriptionally active regions of the genome from intergenic, intronic and genic regions. Signal to background ratios were determined by first calling probes that fluoresced at intensities greater than 99% of the random probes’ signal intensities; therefore, only 1% of fluorescing experimental probes should be false positives.
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| Overall design |
We conducted two-color competitive hybridizations that measure differential expression from three replicates, each using RNA from independent biological extractions. Transcriptional active regions (TARs) were defined by stringing together overlapping probes showing fluorescence above a 1% false positive rate (FPR). Positive probes were joined into a TAR if they were adjacent (maxgap=0, no intermittent non-positive probe) and a TAR’s length had to be at least 45 bp (minrun=45, mid-point first positive probe to mid-point last positive probe, resulting in at least 3 adjacent positive probes for a TAR). The data analysis to measure differential expression of genes and of unannotated TARs was performed using the statistical software package R and Bioconductor with additions and modifications. The signal distributions across chips, samples and replicates were adjusted to be equal according to the mean fluorescence of the random probes on each array. All probes including random probes were quantile-normalized across replicates. Expression-level scores were assigned for each predicted gene based on the median log2 fluorescence over background intensity of probes falling within the exon boundaries.
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| Contributor(s) |
Colbourne JK, Lopez J, Choi J |
| Citation(s) |
21292972 |
| |
| Submission date |
Dec 04, 2010 |
| Last update date |
Jun 25, 2012 |
| Contact name |
Jacqueline Lopez |
| E-mail(s) |
jacqueline.ann.lopez@gmail.com
|
| Phone |
574-631-1902
|
| Organization name |
University of Notre Dame
|
| Department |
Biology
|
| Lab |
Pfrender Laboratory
|
| Street address |
019 Galvin Life Science Center
|
| City |
Notre Dame |
| State/province |
Indiana |
| ZIP/Postal code |
46556 |
| Country |
USA |
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| Platforms (2) |
| GPL11200 |
NimbleGen Daphnia pulex Whole-genome Tiling Array Set (1 of 2) |
| GPL11201 |
NimbleGen Daphnia pulex Whole-genome Tiling Array Set (2 of 2) |
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| Samples (6)
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| GSM634878 |
Sample1_chip1_experimentB_replicate1 |
| GSM634879 |
Sample2_chip1_experimentB_replicate2 |
| GSM634880 |
Sample3_chip1_experimentB_replicate3 |
| GSM634881 |
Sample1_chip2_experimentB_replicate1 |
| GSM634882 |
Sample2_chip2_experimentB_replicate2 |
| GSM634883 |
Sample3_chip2_experimentB_replicate3 |
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| This SubSeries is part of SuperSeries: |
| GSE25823 |
The Ecoresponsive Genome of Daphnia pulex: Empirical annotation of the Daphnia pulex genome |
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| Relations |
| BioProject |
PRJNA142495 |
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