Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Empirical Annotation of the Daphnia pulex genome; Experiment A

ABSTRACT: 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. 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.

ORGANISM(S): Daphnia pulex

SUBMITTER: Jacqueline Lopez

PROVIDER: E-GEOD-25855 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

The ecoresponsive genome of Daphnia pulex.

Colbourne John K JK Pfrender Michael E ME Gilbert Donald D Thomas W Kelley WK Tucker Abraham A Oakley Todd H TH Tokishita Shinichi S Aerts Andrea A Arnold Georg J GJ Basu Malay Kumar MK Bauer Darren J DJ Cáceres Carla E CE Carmel Liran L Casola Claudio C Choi Jeong-Hyeon JH Detter John C JC Dong Qunfeng Q Dusheyko Serge S Eads Brian D BD Fröhlich Thomas T Geiler-Samerotte Kerry A KA Gerlach Daniel D Hatcher Phil P Jogdeo Sanjuro S Krijgsveld Jeroen J Kriventseva Evgenia V EV Kültz Dietmar D Laforsch Christian C Lindquist Erika E Lopez Jacqueline J Manak J Robert JR Muller Jean J Pangilinan Jasmyn J Patwardhan Rupali P RP Pitluck Samuel S Pritham Ellen J EJ Rechtsteiner Andreas A Rho Mina M Rogozin Igor B IB Sakarya Onur O Salamov Asaf A Schaack Sarah S Shapiro Harris H Shiga Yasuhiro Y Skalitzky Courtney C Smith Zachary Z Souvorov Alexander A Sung Way W Tang Zuojian Z Tsuchiya Dai D Tu Hank H Vos Harmjan H Wang Mei M Wolf Yuri I YI Yamagata Hideo H Yamada Takuji T Ye Yuzhen Y Shaw Joseph R JR Andrews Justen J Crease Teresa J TJ Tang Haixu H Lucas Susan M SM Robertson Hugh M HM Bork Peer P Koonin Eugene V EV Zdobnov Evgeny M EM Grigoriev Igor V IV Lynch Michael M Boore Jeffrey L JL

Science (New York, N.Y.) 20110201 6017

We describe the draft genome of the microcrustacean Daphnia pulex, which is only 200 megabases and contains at least 30,907 genes. The high gene count is a consequence of an elevated rate of gene duplication resulting in tandem gene clusters. More than a third of Daphnia's genes have no detectable homologs in any other available proteome, and the most amplified gene families are specific to the Daphnia lineage. The coexpansion of gene families interacting within metabolic pathways suggests that ...[more]

PMID: 21292972

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Project description: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. 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). Transcriptional active regions (TARs) were defined by stringing together overlapping probes showing fluorescence above a 1% false positive rate (FPR). 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.

Dataset Information

Empirical Annotation of the Daphnia pulex genome; Experiment A

Publications

The ecoresponsive genome of Daphnia pulex.

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