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Status |
Public on Jul 10, 2012 |
Title |
PolyA RNA_A. queenslandica precompetent larvae_library2_b |
Sample type |
SRA |
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|
Source name |
Precompetent larvae
|
Organism |
Amphimedon queenslandica |
Characteristics |
developmental stage: Precompetent larvae tissue: Amphimedon tissues
|
Growth protocol |
Amphimedon tissues were collected at different developmental stages: precompetent (<3 hours after emergence from brood chambers); competent (>6 hours after emergence); postlarva (>4 hours after settlement); adult (no brood chambers)
|
Extracted molecule |
polyA RNA |
Extraction protocol |
RNA from larvae and postlarvae was extracted directly with Trizol [Invitrogen]. Adult tissues without brood chambers were cleaned of macroscopic debris then ground in liquid nitrogen before RNA extraction with Trizol. Contaminating DNA was removed using the DNAfree kit [Ambion]. The poly(A) RNA fraction was enriched using the MicroPoly(A)Purist kit [Ambion] and ribosomal RNA was depleted using the RiboMinus Eukaryote kit [Invitrogen]. RNA quality was monitored using the Agilent Bioanalyzer RNA 6000 Pico Assay. Fragment libraries were prepared as described in Cloonan et al. (2008) Nat. Methods 5(7). Briefly, approximately 250ng of purified poly(A) RNA was subjected to 95°C until most of the RNA formed 50-200nt fragments. First-strand cDNA synthesis was primed with a 3â adapter-tagged random hexamer primer using SuperScript II reverse transcriptase. Second strand cDNA was synthesized using a 5â template switching adapter-tagged oligonucleotide. cDNA libraries were amplified using limited PCR cycles and fragments averaging 150bp in length were purified. Libraries were quantified by PCR and fragment size was verified using the Agilent Bioanalyzer DNA 1000 Assay. Fragment libraries were sequenced at 50 base pair reads using the Applied Biosystems SOLiD sequencer (version 3.0).
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Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
AB SOLiD System 3.0 |
|
|
Description |
PRE2 Precompetent larvae (<3 hours after emergence from brood chambers)
|
Data processing |
50nt reads obtained from sequencing runs were analyzed in color space using the SOLiD RNA Analysis Pipeline Tool (http://solidsoftwaretools.com/gf/project/transcriptome). Reads were aligned to sponge contigs using the âanchor-extendâ method as previously described (Tang, Barbacioru et al. (2010) Nature Protocols 5(3)). Assembled sponge contigs and gene models (processed data file build: Aqu1) are publicly available at http://spongezome.metazome.net/ (Srivastava, Simakov et al. (2010) Nature 466(7307)). Peak files were generated by plotting read coverage across all contigs for each run.
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|
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Submission date |
Jun 15, 2011 |
Last update date |
May 15, 2019 |
Contact name |
Cecilia Conaco |
E-mail(s) |
conaco@lifesci.ucsb.edu
|
Phone |
(805)8934586
|
Organization name |
University of California at Santa Barbara
|
Department |
Neuroscience Research Institute
|
Street address |
BioII
|
City |
Santa Barbara |
State/province |
California |
ZIP/Postal code |
93106 |
Country |
USA |
|
|
Platform ID |
GPL13724 |
Series (1) |
GSE29978 |
Transcriptome of the demosponge Amphimedon queenslandica at life cycle transitions |
|
Relations |
SRA |
SRX079850 |
BioSample |
SAMN00630450 |