|
| Status |
Public on Oct 12, 2022 |
| Title |
EpiCs tcf21-/- 96 hpf [Epi-Mut_1] |
| Sample type |
SRA |
| |
|
| Source name |
Sorted epicardial cells
|
| Organism |
Danio rerio |
| Characteristics |
strain: Tg(tcf21:nls-EGFP)+
genotype: tcf21-/-
developmental stage: 96 hpf
cell type: Epicardial cells
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Hearts from 96 hpf Tg(myl7:mcherry), TgBAC(tcf21:nls-EGFP) tcf21+/+ and tcf21-/- zebrafish larvae were manually dissected using forceps. Hearts were dissociated into single-cell suspensions and FACS was performed to sort cardiomyocytes and epicardial cells. Approximately 6000-1000 cells per replicate were used, and total RNA was isolated using the miRNeasy micro kit (Qiagen), combined with on-column DNase digestion.
4 ng of total RNA was used as input for SMART-Seq® v4 Ultra® Low Input RNA Kit (Takara Clontech) for cDNA pre-amplification. Obtained full length cDNA was checked on LabChip and fragmented by Ultrasonication by E220 machine (Covaris). Final Library Preparation was performed by Low Input Library Prep Kit v2 (Takara Clontech).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
Epi-Mut_1
counts.matrix.norm_anno.txt
|
| Data processing |
Trimmomatic version 0.39 was used to trim reads with a quality drop below a mean of Q20 in a window of 10 nucleotides. Only reads between 30 and 150 nucleotides were used in subsequent analyses.
Trimmed and filtered reads were aligned versus the Ensembl Zebrafish genome version DanRer11 (Ensembl release 99) using STAR 2.7.3a with the parameter “outFilterMismatchNoverLmax 0.1” to increase the maximum ratio of mismatches to mapped length to 10%.
The number of reads aligning to genes was counted with featureCounts 1.6.5 tool from the Subread package (Liao et al., 2014). Only reads mapping at least partially inside exons were admitted and aggregated per gene, while reads overlapping multiple genes or aligning to multiple regions were excluded from further analyses.
Differentially expressed genes were identified using DESeq2 version 1.26.0 (Love et al., 2014). To remove a batch effect from the comparison, the replicates were presented to DESeq2 as covariates (DMSO_1/Inhib_1 = 1, DMSO_2/Inhib_2 = 2).
The Ensembl annotation was enriched with UniProt data (release 06.06.2014) based on Ensembl gene identifiers.
Genome_build: DanRer11 (GRCz11)
Supplementary_files_format_and_content: counts.matrix.norm_anno.txt: Tab-delimited text file includes library size normalized counts.
|
| |
|
| Submission date |
May 16, 2021 |
| Last update date |
Oct 12, 2022 |
| Contact name |
Stefan Günther |
| E-mail(s) |
stefan.guenther@mpi-bn.mpg.de
|
| Organization name |
MPI for heart and lung research
|
| Street address |
ludwigtr. 43
|
| City |
bad nauheim |
| ZIP/Postal code |
61231 |
| Country |
Germany |
| |
|
| Platform ID |
GPL20828 |
| Series (1) |
| GSE174505 |
Identification of Tcf21 downstream genes in the epicardial cells and cardiomyocytes by transcriptomic analysis |
|
| Relations |
| BioSample |
SAMN19227675 |
| SRA |
SRX10902993 |
