|
| Status |
Public on May 10, 2017 |
| Title |
e9.5 AVC Hand2 null-mutant (3) |
| Sample type |
SRA |
| |
|
| Source name |
e9.5 AVC Hand2 null-mutant
|
| Organism |
Mus musculus |
| Characteristics |
stain background: Outbred NMRI mice
genotype/variation: Hand2 d/d (null-mutant)
developmental stage: gestational day e9.0-e9.25
tissue: embryonic heart
tissue: Atrioventricular canal tissue
library type: Unstranded
|
| Growth protocol |
The Hand2 d/d (null-Mutant) allele was generated by Cre-mediated deletion of the entire Hand2 coding regions in the germline (PMID: 20386744). Noon of the day the vaginal plug was observed was considered as embryonic day 0.5 (E0.5). Mice and embryos were genotyped by PCR using genomic DNA extracted from toe biopsies and yolk sacs, respectively.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Atrioventricular canals were hand-dissected from isolated hearts using Tungsten needles, flash-frozen individually in Qiagen RTL buffer and stored at -80 °C until genotyped. For each genotype, four replicates of four pooled AVCs with the same distribution of somite stages and sex ratios were prepared and RNA was extracted using Qiagen RNeasy Micro kit according to the manufacturer's protocol."
The quality of total RNA (30-60ng) from the four wild-type and Hand2-deficient samples was analyzed using the RNA 6000 Pico kit in combination with the Agilent 2100 Bioanalyzer. Seven of the eight samples had RIN values >=8.5, while the RNA extracted from one wild-type sample was of suboptimal quality. Therefore, only three wild-type and all four Hand2-deficient samples were processed for sequencing. Ribosomal RNA was depleted using the Ribo-Zero Magnetic Gold Kit (Epicentre). RNA-Seq libraries were prepared using the Clontech SMARTer Low Input RNA kit (Clontech). The resulting cDNA was subjected to eight cycles of PCR amplification and sequenced on a Illumina HiSeq 3000 instrument with a SR50 protocol.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 3000 |
| |
|
| Description |
e9.5_AVC_Hand2_null-mutant_rep3
|
| Data processing |
Reads were aligned to the mm9 reference genome and to the Mus Musculus transcriptome (iGenome refGene) using TopHat v2.0.13 (PMID: 23618408). The option --no-coverage-search was specified, while all the other parameters were left to default.
Gene-wise counts were calculated using htseq-count (PMID: 25260700) setting -s to no and considering the same reference transcriptome. Only those reads with a unique match to the genome were considered (samtools view -q1).
Genome_build: mm9 (NCBI Build 37), transcripts from Mus Musculus Ensembl Build 63.
Supplementary_files_format_and_content: *.counts.txt list the raw counts per gene as inferred by htseq-count.
|
| |
|
| Submission date |
Jan 27, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Iros Barozzi |
| E-mail(s) |
iros.barozzi@meduniwien.ac.at
|
| Organization name |
Medical University Vienna
|
| Street address |
Borschkegasse 8a
|
| City |
Vienna |
| ZIP/Postal code |
1090 |
| Country |
Austria |
| |
|
| Platform ID |
GPL21493 |
| Series (1) |
| GSE94246 |
HAND2 Target Gene Regulatory Networks Control Cardiac Valve Development |
|
| Relations |
| BioSample |
SAMN06277534 |
| SRA |
SRX2524487 |
