Differentially expressed genes in zebrafish vascular development
ABSTRACT: The goal of this experiment was to identify using tissue specific transgenic zebrafish (Fli:EGFP), genes of interest important for embryonic vascular development. Keywords: time course, GFP cell isolation, zebrafish Overall design: The experiment contained 6 samples. A18, A24 = total RNA isolated from 18 and 24 hpf zebrafish embryo; B2, C and D2=RNA isolated from GFP positive cells sorted at 18, 24 and 28 hpf respectively from Tg(Fli1:EGFP) embryos; E2 = RNA isolated from GFP positive cells sorted at 18 hpf from Tg(Flk:GRCFP) embryos.
Project description:The goal of this experiment was to identify using tissue specific transgenic zebrafish (Fli:EGFP), genes of interest important for embryonic vascular development. Keywords: time course, GFP cell isolation, zebrafish The experiment contained 6 samples. A18, A24 = total RNA isolated from 18 and 24 hpf zebrafish embryo; B2, C and D2=RNA isolated from GFP positive cells sorted at 18, 24 and 28 hpf respectively from Tg(Fli1:EGFP) embryos; E2 = RNA isolated from GFP positive cells sorted at 18 hpf from Tg(Flk:GRCFP) embryos.
Project description:The WT and Tg(RE:HSE:EGFP) F2 generation embryos were heated shocked at 38°C for 1 hour at 16 hpf, then raised to 3 dpf. Total RNAs were isolated with Trizol (Invitrogen). The samples were processed and subsequently analyzed in triplicate on Zebrafish Oligo Microarrays (Agilent Technologies Italia, Italy) which contain 43,554 sets of probes. The microarrays were scanned in an Agilent DNA Microarray Scanner and the images were processed using Feature Extraction software to identify changes of gene expression signatures in Tg(RE:HSE:EGFP) zebrafish embryos. Overall design: RUNX1-Evi-1 fusion gene was induced into wild type AB line and measured at 72 hours after 1hr heat-shock at 38°C at 16 hpf. Wild type AB line was used as the control.
Project description:Purpose: RNA seq analysis from isolated zebrafish HSPCs at 48 hpf from control morpholino and dnmt3bb.1 morpholino injected embryos Overall design: HSPCs were isolated by FACS sorting using transgenic line Tg(kdrl:mApple, cmyb:GFP) as described by Bertrand et al 2010 Total RNA from isolated HSPCs at 48hpf
Project description:In order to discover the targets of Foxj1, we made transgenic zebrafish in which Foxj1 is ubiquitously overexpressed in response to heat [Tg(hsp70::foxj1a)]. Transgenic embryos and wild type control embryos were collected, given two heat shocks (at 18 hours post fertilization (hpf) and 20 hpf), then analyzed at 22 hpf. Gene expression profiles of embryos overexpressing Foxj1a were compared to gene expression profiles of wild type embryos using Nimblegen whole transcriptome zebrafish microarrays.
Project description:Purpose: To identify genes that are transcriptionally controlled by Notch signaling during zebrafish lateral line proneuromast formation. Methods: We isolated primordium cells from dissected tails of 36 hpf Tg((cldnB:GFP);Tg(cldnB:gal4) x Tg(UAS:nicd)) and sibling Tg((cldnB:GFP);Tg(cldnB:gal4)) embryos by FACS and performed RNASeq analysis. Results: Using an optimized data analysis workflow, we mapped about 26 million sequence reads per sample to the zebrafish genome (build danRer10) and identified 32,105 transcripts in the dissociated tails of WT and NICD zebrafish with TopHat workflow. Approximately 2% of the transcripts showed differential expression between the WT and NICD tails, with a fold change ≥0.5 and p value <0.01. Conclusion: RNASeq analyses revealed that Notch signaling cell-autonomously induces apical constriction and cell adhesion. Overall design: Zebrafish lateral line mRNA profiles of 36 hours wild type (WT) and NICD embryos were generated in triplicate, using HiSeq 2500 (Illumina).
Project description:This work sought to identify endothelial-specific enhancer elements by applying ATAC-Seq to nuclei isolated from Tg(fli1a:egfp) transgenic zebrafish embryos. Overall design: for comparison of cell type specific gene expression and ATAC-Seq datasets, we performed parallel RNAseq in duplicate on GFPplus and minus cells isolated from Tg(fli1a:egfp) zebrafish embryos
Project description:We isolated and expression profiled V0v spinal interneurons, as well as all spinal neurons and all trunk cells using Fluorescence Activated Cell Sorting (FACS) of different transgenic zebrafish lines in which these cells express fluorescent proteins at 27 hours post fertilization (hpf). We used a custom-designed Agilent microarray to study the expression of transcription factors in these cell types. Overall design: Twelve samples were analysed in total, all at 27 hours post fertilisation (hpf). Four biological replicates were performed for each of the three cell types under study - V0v spinal interneurons (isolated from Tg(evx1:EGFP)SU1), all spinal neurons (isolated from Tg(elavl3:EGFP)) and all trunk cells (FAC-sorted trunk cells).
Project description:Fluorescence-Activated Nuclei Sorting (FANS)-assisted Assay for Transposase Accessible Chromatin sequencing (ATAC-seq) This work sought to identify endothelial-specific enhancer elements by applying ATAC-Seq to nuclei isolated from Tg(fli1a:egfp) transgenic zebrafish embryos. Overall design: ATAC-Seq performed on GFPplus and GFPminus nuclei isolated from Tg(fli1a:egfp) embryos. Separate ATAC-Seq libraries were constructed from GFPplus and minus nuclei isolated on three separate occasions to give biological triplicate samples and 6 total samples.
Project description:Transcriptome data from zebrafish single and bulk cells from blood and testes. Blood cells were collected from adult Tg(cd4:mCherry), Tg(cd41:EGFP), Tg(gata1a:GFP), Tg(lyz:DsRed2), Tg(mfap4:tdTomato), Tg(mpx:EGFP), Tg(runx1:mCherry), Tg(tal1:EGFP) and Tubingen Long Fin wild type fish.
Project description:In order to detect transcriptional differences between primitive and definitive hematopoietic stem and progenitor cells during regular development in the zebrafish embryo, gata1-GFP+/+(18 somites), lmo2-GFP+/+ (18 somites and 35 hpf)1 and cd41-GFP+/+ (35 hpf) cells from transgenic embryos were individually separated from GFP-/- cells by flow cytometry at the indicated stages. For each individual population, pools of 600 - 1500 transgenic embryos were collected. After RNA extraction, labelled cRNA was hybridized onto Affymetrix microarrays. Individual experiments were performed with 2 or 3 biological replicates. Keywords: cell type comparison Overall design: gata1-GFP+/+(18 somites), lmo2-GFP+/+ (18 somites and 35 hpf)1 and cd41-GFP+/+ (35 hpf)2 cells were separated from GFP-/- cells by flow cytometry at the indicated stages. Sorting was based on propidium iodide exclusion, forward scatter, and GFP fluorescence, using a FACSVantage flow cytometer (Beckton Dickinson). Sorted cell populations were run twice to optimize cell purity. Total RNA from cell-sorted populations was extracted with TRIzol reagent (Invitrogen) and purified on RNeasy resins (Qiagen) according to the manufacturer’s recommendations. Total RNA was subjected to two rounds of linear amplification (Ambion) and hybridized to Affymetrix zebrafish Gene Chips, according to Affymetrix guidelines. After staining with a streptavidin-phycoerythrin conjugate (Molecular Probes), the fluorescence of bound RNA was quantitated by using a Gene Chip scanner (Affymetrix). The raw expression data were calculated and, after pairing of GFP+/+ and GFP-/- samples, statistically analyzed using methods implemented in Bioconductor’s “affy” package and available custom scripts 3,4. references: 1. Zhu, H. et al. Regulation of the lmo2 promoter during hematopoietic and vascular development in zebrafish. Dev Biol 281, 256-69 (2005). 2. Lin, H. F. et al. Analysis of thrombocyte development in CD41-GFP transgenic zebrafish. Blood 106, 3803-10 (2005). 3. Choe, S. E., Boutros, M., Michelson, A. M., Church, G. M. & Halfon, M. S. Preferred analysis methods for Affymetrix GeneChips revealed by a wholly defined control dataset. Genome Biol 6, R16 (2005). 4. Weber, G. J. et al. Mutant-specific gene programs in the zebrafish. Blood 106, 521-30 (2005).