|
| Status |
Public on May 27, 2014 |
| Title |
second_trimester_euploid_male1 [RNA-Seq] |
| Sample type |
SRA |
| |
|
| Source name |
cell-free RNA from second trimester amniotic fluid supernatant
|
| Organism |
Homo sapiens |
| Characteristics |
gender: male
source: second trimester amniotic fluid supernatant
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted from 10 mL amniotic fluid supernatant (centrifugation at 350g, 4C, 10, min).All samples were processed using the Qiagen Circulating Nucleic Acid kit with an on-column DNase digestion step to remove genomic DNA. The RNA was then purified and concentrated with the RNeasy MinElute Clean up kit and eluted in RNasefree water. RNA was converted to cDNA and amplified using the Ovation RNA-Seq System V2 and then purified with the MinElute Reaction Cleanup Kit.
Library preparation was undertaken on 500 ng cDNA according to standard practice with the alterations described: Fragmentation was not undertaken due to the already degraded and fragmented nature of cffRNA. In light of the small amount of starting material, a bead-based purification (AmPure XP Beads, Agencourt, Beckman Coulter, USA) was used in lieu of a gel-based size selection for the purpose of library clean up. Bioanalysis was performed to ensure the absence of primer-dimer peaks as well as determine the approximate insert size. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Illumina Hi-Seq 2000 following the manufacturer's protocols. One paired-end indexed library was sequenced per sample to a length of 50 nucleotides per mate.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
gene expression data from normal second trimester fetus
|
| Data processing |
Basepair calls performed using CASAVA-1.8.2
Reads were aligned to the Hg19 UCSC genome using the Spliced Transcripts Alignment to a Reference (STAR) aligner (version 2.3.0e). The STAR reference genome was created with a specific library of splice junctions (Gencode version 15 from the Ensembl 70 March 2012 freeze) and the sdjbOverhang 49 option. The following custom settings were used in STAR alignment scripts: sjdbScore, outFilterType BySSJout, outFilterIntronMotifs RemoveNoncanonicalUnannotated, outSAMstrandField intronMotif. All other parameters were run according to default including multi-mapping of up to 10.
Cufflinks version 2.1.1 was used with the Hg19 goldenPath UCSC annotation GTF file (Feb 2009 assembly). All default Cufflinks parameters were employed, including multimapping.
Genome_build: UCSC Hg19
Supplementary_files_format_and_content: tab delimited text files of Cufflinks FPKM tracking File output including FPKM values
|
| |
|
| Submission date |
Aug 14, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Lillian Zwemer |
| E-mail(s) |
[email protected]
|
| Organization name |
Tufts Medical Center
|
| Department |
Mother Infant Research Institute
|
| Street address |
800 Washington St, Box 394
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02111 |
| Country |
USA |
| |
|
| Platform ID |
GPL11154 |
| Series (2) |
| GSE49890 |
RNA-Seq and expression microarray highlight different aspects of the fetal amniotic fluid transcriptome [RNA-Seq] |
| GSE49893 |
RNA-Seq and expression microarray highlight different aspects of the fetal amniotic fluid transcriptome |
|
| Relations |
| BioSample |
SAMN02317091 |
| SRA |
SRX335393 |
