Project description:Percellome analysis of whole Xenopus embryos at developmental stage 18 Xenopus embryos were treated with TTNPB (RAR-agonist), AGN193109 (RAR-antagonist), or Control Vehicle at Nieuwkoop and Faber Stage 7. Embryos were collected at stage 18, followed by RNA extraction, hybridization on Affymetrix microarrays, and Percellome analysis.
Project description:The development of massively parallel sequencing technologies has revolutionized transcriptome analysis. Sequencing of total cDNA (RNA-Seq) can determine the expression levels of known and novel transcripts with high sensitivity from various developmental stages or conditions. Here we report a robust method for RNA-Seq in Xenopus laevis and apply it to understanding how modulation of retinoic acid signaling alters the transcriptome in early Xenopus laevis development. Three biological conditions were tested: early blastula stage embryos were treated with 1. a retinoic acid receptor (RAR) agonist NRX204647 (4647) at a concentration that stimulated all 3 RAR subtypes (alpha, beta, and gamma); 2. The same agonist at an RAR[gamma] selective dose, and 3. vehicle control. Five single-clutch replicates were obtained for each chemical treatment group, and harvested at neurula stage. We found that single-clutch replicates reflect the stochastic variation of the general outbred population and contribute more statistical power to RNA-Seq experiments due to their feasibility of replication. Our RNA-Seq dataset identified 1590 up-regulated and 685 down-regulated unique genes differentially regulated by all RAR subtypes, and 160 up-regulated and 60 down-regulated genes likely to be regulated specifically by RAR[gamma]. Differential expression detected by RNA-Seq was validated for selected genes by QPCR, which demonstrated nearly 100% quantitative agreement with the deep sequencing data. We further validated RAR-responsive genes by comparison with two previous, published microarray datasets and found substantial agreement. Gene ontology analysis identified RAR targets which may underlie such developmental processes as axial elongation, neurogenesis/synaptogenesis, dorsoventral regulation of the retina, homeotic fate specification, and central nervous system development. We investigated RAR[gamma]-selective targets identified by RNA-Seq and inferred that transcriptional repression by unliganded RARs is of substantial importance to embryonic patterning events. Overall, this paper demonstrates the utility of RNA-Seq in Xenopus laevis, obviating the perceived requirement to use X. tropicalis for genomic analyses.
Project description:Transcriptional profiling of Xenopus laevis embryos and ectoderm (animal caps) comparing embryos injected with control morpholino with embryos injected with the morpholino mixture PVD2, which knocks down all three Xenopus PouV proteins. Whole embryos (WE) or animal caps (AC) were collected at late blastula (9) or early gastrula (10) stages from Control and PVD2 morphants.
Project description:The aim of the experiment was to identify genome wide binding sites for retinoic acid receptor beta (RARB) in RARB agonist treated human metastatic pancreatic ductal adenocarcinoma cells (SUIT2). Datasets are prsented for the ChIP-seq analysis for SUIT2 cells after 72 h treatment with either DMSO (vehicle control), 1 µM RAR-β agonist (CD 2314, Tocirs 3824), or 1 µM RAR-β antagonist (LE 135, Tocris 2021).