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| Status |
Public on Jun 27, 2016 |
| Title |
Transcriptome-wide profiling of Spadetail-regulated genes in early somite progenitors |
| Organism |
Danio rerio |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
We integrate zebrafish embryology with photoactivatable caged morpholinos (cMO), the photoactivatable lineage tracer caged fluorescein-dextran (cFD), fluorescence-activated cell-sorting (FACS) and RNA sequencing (RNA-seq) to identify Spadetail-regulated genes in mesodermal cells enriched for early somite progenitors.
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| Overall design |
1 to 4-cell stage zebrafish embryos were injected with either cFD alone or in combination with the spt cMO and allowed to develop until 6 hours post-fertilization (hpf). Early ventral somite progenitors were then optically targeted with ultraviolet (360 nm) light using a 100-μm diameter circular diaphragm. Embryos were permitted to develop until 9 hpf, dechorionated, and dissociated into single-cells. Fluorescein-positive cells were then isolated by FACS. Five experimental replicates were performed. Sequencing libraries were then prepared from purified cells according to the CEL-Seq protocol (Hashimshony, T et al. Cell Reports. 2012). In short, total RNA from each sample were individually barcoded and combined prior to sequencing on a HiSeq 2000. Reads were processed and aligned to zebrafish assembly Zv8 using custom CEL-seq scripts within the Galaxy bioinformatics platform. Count files were then analyzed using EdgeR statistical analysis to determine differentially expressed genes.
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| Contributor(s) |
Payumo AY, Chen JK |
| Citation(s) |
27376691 |
| Submission date |
Jul 08, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
James Chen |
| Organization name |
Stanford University
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| Department |
Chemical and Systems Biology
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| Lab |
James Chen
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| Street address |
269 W. Campus Drive, CCSR 3150
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| City |
Stanford |
| State/province |
CA |
| ZIP/Postal code |
94035 |
| Country |
USA |
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| Platforms (1) |
| GPL14875 |
Illumina HiSeq 2000 (Danio rerio) |
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| Samples (10)
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| Relations |
| BioProject |
PRJNA289199 |
| SRA |
SRP060505 |
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