|
Status |
Public on Sep 12, 2011 |
Title |
RNA Capture sequencing 4 (454) |
Sample type |
SRA |
|
|
Source name |
Foot (c45) fibroblasts
|
Organism |
Homo sapiens |
Characteristics |
cell type: Primary foot fibroblasts sample type: RNA Capture sequencing
|
Growth protocol |
Primary human female fetal lung fibroblasts and human male foot fibroblasts in DMEM supplemented with 10% FBS at 37 °C with 5% CO2 as previously described (J. L. Rinn et. al., PLoS Genet, 2006).
|
Extracted molecule |
total RNA |
Extraction protocol |
454 GS-FLX Titanium Sequencing libraries from c45 and FL fibroblast cDNA were constructed using the 454 LifeSciences GSFLX Titanium Kit according to manufacturers instructions. All single-stranded DNA product from library preparation was utilized as template in a Pre-Hybridization Linker Mediated PCR reaction to ensure that the plurality of the molecules contained adapters on both sides of the putative cDNA inserts. The amplified material was recovered with a Qiaquick column and DNA evaluated electrophoretically by an Agilent Bioanalyzer 2100. Library fragment sizes were found to be between 500-700 bp. Samples were purified and combined with C0T-1 DNA, enhancing oligos, NimbleGen hybridization buffer and Component A prior to capture. Hybridisation and post-hybridisation LMPCR were performed with identical conditions. The resulting post capture enriched sequencing libraries were sequenced on 454 Genome Sequencer FLX System using Titanium chemistry.
|
|
|
Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
454 GS FLX Titanium |
|
|
Data processing |
Roche 454 reads were firstlycombined and aligned to the human genome (hg19) using Blat (http://www.soe.ucsc.edu/~kent) with the following non-default parameters; minIdentity=90, minScore=100. Resultant *.psl files were converted into *.sam files using bedTools and samtools.
|
|
|
Submission date |
May 03, 2011 |
Last update date |
May 15, 2019 |
Contact name |
Tim R. Mercer |
Organization name |
Garvan
|
Lab |
Prof. John Mattick
|
Street address |
384 Victoria Street Darlinghurst NSW 2010
|
City |
Sydney |
State/province |
New South Wales |
ZIP/Postal code |
2010 |
Country |
Australia |
|
|
Platform ID |
GPL14603 |
Series (2) |
GSE29040 |
RNA Capture-Seq resolves the deep complexity of the human transcriptome (454) |
GSE29041 |
RNA Capture-Seq resolves the deep complexity of the human transcriptome |
|
Relations |
SRA |
SRX060821 |
BioSample |
SAMN00263081 |