ATACseq of kdrl-GFP+ endothelial cells in 28hpf zebrafish embryos
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PROVIDER: PRJNA601524 | ENA |
REPOSITORIES: ENA
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Project description:The goal of this study is to analyse the open chromatin regions via ATACseq in zebrafish embryonic endothelial cells at 26-27hpf using Tg(Kdrl:GFP) transgenic zebrafish embryos. 40000-50000 cells were FAC-sorted directly into 100 µl of 1x HBSS/10 mM HEPES/0.25% BSA buffer for Tagmentation (as described by Buenrostro et al. (2013), Nat. Methods 10, 1213-1218), but with 1.5 µl Tn5 transposase (Illumina) in a 50 µl reaction volume. DNA was purified using the QIAquick PCR purification kit (Qiagen). Fragments were amplified for 16 cycles.
2020-01-16 | GSE143763 | GEO
Project description:The zebrafish is a powerful model for the study of hematopoietic stem and progenitor cells (HSPC). We have developed a novel HSPC-specific transgenic line (Runx1+23:GFP). We have used this line in time-lapse live imaging studies to track the migration of HSPC during development. We have also performed a chemical genetic screen to find small molecules that modulate HSPC numbers during development. Treating embryos from 2-3 days post fertilization (2-3 dpf) then fixing for in situ staining with HSPC probes cmyb and runx1, we found the compound lycorine increased HSPC numbers. Applying this compound during time-lapse live imaging showed increased accumulation of Runx+ HSPC in the caudal hematopoietic tissue (CHT). Treatment from 2-3 dpf, then washing off the compound, had a sustained effect on the size of the HSPC with Runx+ numbers higher at 5 and 7 dpf. We have performed microarray analysis to elucidate the molecular changes within HSPC and endothelial cells after Lycorine treatment. We treated Runx1+23:GFP;kdrl:DsRed2 embryos from 2-3 dpf with 75 uM lycorine in 1% DMSO. We then dissociated the embryos and sorted the Runx+ GFP cells, the kdrl+ DsRed2 cells, and the non-fluorescent negative cells from the total embryo as a comparator population. Total RNA was amplified and biotin labled for hybridization on Affymetrix microarrays. 18 samples were collected and analyzed. There are 3 biological replicates. There are 3 cell type populations: 1) Runx+ HSPC; 2) kdrl+ endothelial cells; 3) non-fluorescent negative cells. There are cell populations from dissociated Lycorine-treated embryo pools, and control DMSO-treated embryo pools.
2015-01-15 | E-GEOD-56015 | biostudies-arrayexpress
Project description:The goal of this study is to compare the NGS-derived heart transcriptome profiling (RNA-seq) between kdrl GFP positive endothelial cells of 4-HT induced Tg(ubi:loxp-DsRed-stop-loxp-DN-xBrg1; kdrl:CreER; kdrl:eGFP) (DNK) and Tg(ubi:loxp-DsRed-stop-loxp-DN-xBrg1; kdrl:eGFP) (CtrlK) injured hearts.
2022-07-09 | GSE154193 | GEO
Project description:We profiled the gene expression patterns of undisturbed endothelial cells in living animals using a novel ‘AngioTag’ zebrafish transgenic line that permits isolation of actively translating mRNAs from endothelial cells in their native environment. This transgenic line uses the endothelial cell-specific kdrl promoter to drive expression of an epitope tagged Rpl10a 60S ribosomal subunit protein, allowing for Translating Ribosome Affinity Purification (TRAP) of actively translating endothelial cell mRNAs. We also collected the whole embryo translatome using the TRAP protocol and a ubiquitously expressed tagged Rpl10a, and the endothelial transcritome by collecting the GFP+ endothelial cells that had the kdrl promoter driving GFP.
2019-03-29 | GSE121973 | GEO
Project description:The zebrafish is a powerful model for the study of hematopoietic stem and progenitor cells (HSPC). We have developed a novel HSPC-specific transgenic line (Runx1+23:GFP). We have used this line in time-lapse live imaging studies to track the migration of HSPC during development. We have also performed a chemical genetic screen to find small molecules that modulate HSPC numbers during development. Treating embryos from 2-3 days post fertilization (2-3 dpf) then fixing for in situ staining with HSPC probes cmyb and runx1, we found the compound lycorine increased HSPC numbers. Applying this compound during time-lapse live imaging showed increased accumulation of Runx+ HSPC in the caudal hematopoietic tissue (CHT). Treatment from 2-3 dpf, then washing off the compound, had a sustained effect on the size of the HSPC with Runx+ numbers higher at 5 and 7 dpf. We have performed microarray analysis to elucidate the molecular changes within HSPC and endothelial cells after Lycorine treatment. We treated Runx1+23:GFP;kdrl:DsRed2 embryos from 2-3 dpf with 75 uM lycorine in 1% DMSO. We then dissociated the embryos and sorted the Runx+ GFP cells, the kdrl+ DsRed2 cells, and the non-fluorescent negative cells from the total embryo as a comparator population. Total RNA was amplified and biotin labled for hybridization on Affymetrix microarrays.
2015-01-15 | GSE56015 | GEO
Project description:The goal of this study is to analyse the open chromatin regions via ATACseq in different sub-types of the zebrafish embryonic dorsal aorta (DA) at 28-30 hpf using TgBAC(runx1P2:Citrine);Tg(kdrl:mCherry) double-transgenic zebrafish embryos. 2000-3000 cells were FAC-sorted directly into 100 µl of 1x HBSS/10 mM HEPES/0.25% BSA buffer for Tagmentation (as described by Buenrostro et al. (2013), Nat. Methods 10, 1213-1218), but with 1.5 µl Tn5 transposase (Illumina) in a 50 µl reaction volume. DNA was purified using the QIAquick PCR purification kit (Qiagen). Fragments were amplified for 16 cycles. Genomic control DNA was isolated from 30.000 cells using the DNeasy Blood & Tissue kit (Qiagen) and eluted with EB buffer. Tagmentation was performed as described. Fragments were amplified for 9 cycles. Sequencing was performed on a NextSeq machine. Sequenced reads were checked for base qualities, trimmed where 20% of the bases were below quality score 20, and filtered to exclude adapters using Trimmomatic (Version 0.32).
2019-06-06 | GSE132258 | GEO
Project description:Microarray analysis of triplicate RNA samples isolated from kdrl:eGFP-sorted ECs of wildtype, npas4l-/-, etsrp-/-, and sox32-/- zebrafish embryos in Tg(kdrl:EGFP) transgenic background between 18 and 18.5 hpf.
2020-10-20 | E-MTAB-9698 | biostudies-arrayexpress
Project description:To decipher the molecular action of endothelial Brg1 in the regulation of zebrafish heart regeneration, we applied Tg(kdrl:eGFP) to label cardiac endothelial cells including the endocardium, and achieved endothelium-specific over-expression of DN-xBrg1 by using the compound zebrafish line consisting of Tg(ubi:loxp-DsRed-STOP-loxp-DN-xBrg1; kdrl:CreER; kdrl:eGFP) (defined as DNK), while we used Tg(ubi:loxp-DsRed-STOP-loxp-DN-xBrg1; kdrl-eGFP) as control (CtrlK) in the presence of 4-HT starting at 3 days before ventricular resection. The kdrl:eGFP endothelial cells, which were sorted by FACS (fluorescence-activated cell sorting) from CtrlK and DNK hearts at 7 dpa, were subjected to RNA-seq analysis.
2023-04-03 | GSE200936 | GEO
Project description:We report gene expression data for FACS sorted zebrafish kdrl(flk1):GFP + and kdrl:mCherry+;sele(5.3kb):GFP+ double positive cells collected from whole embryos at 72 hours post fertilization (hpf). We also report gene expression data for the kdrl:GFP negative fraction.
2023-03-23 | GSE124148 | GEO
Project description:Fish, JE, Santoro, MM, Morton, SU, Yu, S, Yeh, RF, Wythe, JD, Ivey, KI, Bruneau, BG, Stainier, DYR, and Srivastava, D. (2008). miR-126 Regulates Angiogenic Signaling and Vascular Integrity. Developmental Cell 15, 272-284. Precise regulation of the formation, maintenance, and remodeling of the vasculature is required for normal development, tissue response to injury, and tumor progression. How specific microRNAs intersect with and modulate angiogenic signaling cascades is unknown. Here, we identified microRNAs that were enriched in endothelial cells derived from mouse embryonic stem (ES) cells and in developing mouse embryos. We found that miR-126 regulated the response of endothelial cells to VEGF. Additionally, knockdown of miR-126 in zebrafish resulted in loss of vascular integrity and hemorrhage during embryonic development. miR-126 functioned in part by directly repressing negative regulators of the VEGF pathway, including the Sprouty-related protein SPRED1 and phosphoinositol-3 kinase regulatory subunit 2 (PIK3R2/p85-β). Increased expression of Spred1 or inhibition of VEGF signaling in zebrafish resulted in defects similar to miR-126 knockdown. These findings illustrate that a single miRNA can regulate vascular integrity and angiogenesis, providing a new target for modulating vascular formation and function. Experiment Overall Design: To identify the genes regulated by miR-126 in vivo, flk1:GFP transgenic zebrafish embryos (which express GFP in the endothelium) were injected at the one-cell stage with 4 ng of two independent antisense morpholinos that block the processing of the miR-126 pri-cursor. At 48 hours post-fertilization, endothelial cells were isolated by fluorescence-activated cell sorting from flk1:GFP transgenic fish, and RNA was extracted. Arrays were performed on four biological replicates of control and two independent miR-126 morpholinos.
2008-10-25 | E-GEOD-12012 | biostudies-arrayexpress