DOI: 10.5061/dryad.v5r6077
title: Data from: Stem cell differentiation trajectories in Hydra resolved at single-cell resolution

description: The adult Hydra polyp continually renews all of its cells using three
 separate stem cell populations, but the genetic pathways enabling this
 homeostatic tissue maintenance are not well understood. We sequenced
 24,985 Hydra single-cell transcriptomes and identified the molecular
 signatures of a broad spectrum of cell states, from stem cells to
 terminally differentiated cells. We constructed differentiation
 trajectories for each cell lineage and identified gene modules and
 putative regulators expressed along these trajectories, thus creating a
 comprehensive molecular map of all developmental lineages in the adult
 animal. In addition, we built a gene expression map of the Hydra nervous
 system. Our work constitutes a resource for addressing questions regarding
 the evolution of metazoan developmental processes and nervous system
 function.
description: Hydra_URD_analysisURD analysis files describing the reconstruction of differentiation trajectories for various cell populations (also available as knitted pdfs). Original simulation files needed to reproduce the original analysis can be found in archive Hydra_URD_cluster_results. This archive also includes a tutorial how to visualize gene expression on trajectories.Hydra_URD_cluster_resultsSimulations results needed to reproduce the published study exactly. For information on how these objects were used/generated see analysis files in archive Hydra_URD_analysis.Hydra_URD_analysis_objectsURD objects (which contain tSNE projections for all tissues and trees for complex tissues) and spline objects (which contain smoothed curves that represent gene expression along particular trajectories).Hydra_Seurat_NMF_regulators_analysesAnalysis files summarizing cell clustering, cell subclustering, cluster annotation, data exploration, non-negative matrix factorization (NMF) analysis and identification of candidate regulators of cell stateHydra_Seurat_EctoSeurat (2.3.4) object. Subcluster analysis for ectodermal epithelial cells. The analysis that resulted in this object is outlined in SA03_SubclustEpithelialCells.Rmd in archive "Hydra_Seurat_NMF_regulators_analyses". The subset of cells in this object was the starting point for URD trajectory reconstruction.Hydra_Seurat_Endo_lineage_plotSeurat (2.3.4) object. Subcluster analysis for endodermal epithelial cells. The analysis that resulted in this object is outlined in SA03_SubclustEpithelialCells.Rmd in archive Hydra_Seurat_NMF_regulators_analyses. The subset of cells and the tSNE representation in this object were used to visualize endodermal epithelial gene expression.Hydra_Seurat_EndoSeurat (2.3.4) object. Subcluster analysis for endodermal epithelial cells. The analysis that resulted in this object is outlined in SA03_SubclustEpithelialCells.Rmd in archive Hydra_Seurat_NMF_regulators_analyses. The subset of cells in this object was the starting point for URD trajectory reconstruction.Hydra_Seurat_NeuronsSeurat (2.3.4) object. Subcluster analysis for neuronal cells and placement of neurons. The analysis that resulted in this object is outlined in SA05_SubclustNeuronalCells.Rmd in archive Hydra_Seurat_NMF_regulators_analyses.nmfInputs and results of non-negative matrix factorization (NMF) analyses for various subsets of cells. We make use of NMF results throughout the analyses (see analysis files in archives Hydra_Seurat_NMF_regulators_analyses and Hydra_URD_analysis). These analyses expect directory nmf/ to be placed in the same location as the analysis markdowns (Rmd files). A sample NMF analysis (enK40) is presented in document SA07_NMF.Rmd available in archive Hydra_Seurat_NMF_regulators_analyses.Candidate_RegulatorsThis archive contains material to reproduce the identification of candidate regulators of cell states. The analysis is presented in SA08_MotifEnrichmentAnalysis.Rmd in archive Hydra_Seurat_NMF_regulators_analyses. The analysis document holds more information on the objects that can be found in this archive.Drop_seq_referencesThis archive contains fasta files for the genome (Hydra 2.0 assembly available at https://research.nhgri.nih.gov/hydra/) and the de novo Hydra vulgaris (AEP) transcriptome (aepLRv2) as well as scripts to create bowtie2 references. It also holds .dict and .refFlat files used by the Drop-seq pipeline.Hydra_Seurat_ICSeurat (2.3.4) object. Subcluster analysis for interstitial cells. The analysis that resulted in this object is outlined in SA04_SubclustInterstitialCells.Rmd in archive Hydra_Seurat_NMF_regulators_analyses. The subset of cells in this object was the starting point for URD trajectory reconstruction.Hydra_Seurat_Whole_GenomeSeurat (2.3.4) object. Cluster analysis after mapping to the Hydra 2.0 genome reference (available at at https://research.nhgri.nih.gov/hydra/). The analysis that resulted in this object is outlined in SA06_ClustGenome.Rmd in archive Hydra_Seurat_NMF_regulators_analyses.Hydra_Seurat_Whole_TranscriptomeSeurat (2.3.4) object. Cluster analysis after mapping to the Hydra transcriptome reference (aepLRv2). The analysis that resulted in this object is outlined in SA02_ClustTranscriptome.Rmd in archive Hydra_Seurat_NMF_regulators_analyses.Drop-seq_pipeline_initialQC.tarDrop-seq pipeline scripts and initial cell quality control. The raw reads (input into the Drop-seq pipeline) and the processed count matrix (output of the initial cell QC) are archived at NCBI GEO (accession number GSE121617).transcriptome_aepLRv2.tarGeneration of a de novo low redundancy transcriptome (aepLRv2) for Hydra vulgaris strain AEP and reference annotation.