too-many-cells

Table of Contents

Website

See https://github.com/GregorySchwartz/too-many-cells for latest version.

See the bioRxiv paper for more information about the algorithm.

pruned_tree.png

Description

too-many-cells is a suite of tools, algorithms, and visualizations focusing on the relationships between cell clades. This includes new ways of clustering, plotting, choosing differential expression comparisons, and more! While too-many-cells was intended for single cell RNA-seq, any abundance data in any domain can be used. Rather than opt for a unique positioning of each cell using dimensionality reduction approaches like t-SNE, UMAP, and PCA, too-many-cells recursively divides cells into clusters and relates clusters rather than individual cells. In fact, by recursively dividing until further dividing would be considered noise or random partitioning, we can eliminate noisy relationships at the fine-grain level. The resulting binary tree serves as a basis for a different perspective of single cells, using our birch-beer visualization and tree measures to describe simultaneously large and small populations, without additional parameters or runs. See below for a full list of features.

New features for current version

  • A new R wrapper was written to quickly get data to and from too-many-cells from R. Check it out here!
  • Now works with Cellranger 3.0 matrices in addition to Cellranger 2.0
  • Can prune (make into leaves) specified nodes with --custom-cut.
  • Can analyze sets of features averaged together (e.g. gene sets). Breaks API, so update your --draw-leaf "DrawItem (DrawContinuous \"Cd4\")" argument to --draw-leaf "DrawItem (DrawContinuous [\"Cd4\"])" (notice the list notation).
  • Outputs values from differential entry point plots (from --genes), and can aggregate genes by average.

Installation

We provide multiple ways to install too-many-cells. We recommend installing stack (see below), but we also have docker images and a Dockerfile to use in any system in case you have a custom build (for instance, a non-standard R installation) or difficulty installing. macOS and Windows users: too-many-cells was built and tested on linux, so we highly recommend using the docker image (which a completely isolated environment which requires no compiling or installation, other than docker itself) as there may be difficulties in installing the dependencies. There are, however, additional instructions for macOS here if you really want to compile it.

Dependencies

You may require the following dependencies to build and run (from Ubuntu 14.04, use the appropriate packages from your distribution of choice):

  • build-essential
  • libgmp-dev
  • libblas-dev
  • liblapack-dev
  • libgsl-dev
  • libgtk2.0-dev
  • libcairo2-dev
  • libpango1.0-dev
  • graphviz
  • r-base
  • r-base-dev

To install them, in Ubuntu:

sudo apt install build-essential libgmp-dev libblas-dev liblapack-dev libgsl-dev libgtk2.0-dev libcairo2-dev libpango1.0-dev graphviz r-base r-base-dev

too-many-cells also uses the following packages from R:

  • cowplot
  • ggplot2
  • edgeR
  • jsonlite

To install them in R,

install.packages(c("ggplot2", "cowplot", "jsonlite"))
install.packages("BiocManager")
BiocManager::install("edgeR")

Install stack

See https://docs.haskellstack.org/en/stable/README/ for more details.

curl -sSL https://get.haskellstack.org/ | sh
stack setup

Install too-many-cells

Source

Probably the easiest method if you don't want to mess with dependencies (outside of the ones above).

git clone https://github.com/GregorySchwartz/too-many-cells.git
cd too-many-cells
stack install

Online

We only require stack (or cabal), you do not need to download any source code (but you might need the stack.yaml dependency versions), just run the following command to place too-many-cells in your ~/.local/bin/:

stack install too-many-cells

If you run into errors like Error: While constructing the build plan, the following exceptions were encountered:, then follow it's advice. Usually you just need to follow the suggestion and add the dependencies to the specified file. For a quick yaml configuration, refer to https://github.com/GregorySchwartz/too-many-cells/blob/master/stack.yaml. Relies on eigen-3.3.4.1 right now.

Docker

Different computers have different setups, operating systems, and repositories. Do put the entire program in a container to bypass difficulties (with the other methods above), we user docker. So first, install docker.

To get too-many-cells (replace 0.1.5.0 with any version needed):

docker pull gregoryschwartz/too-many-cells:0.1.5.0

To run too-many-cells in a docker container:

sudo docker run gregoryschwartz/too-many-cells:0.1.5.0 -h

Docker won't be able to find your files by default. You need to mount the folders with -v in order to have docker read and write from and to the filesystem, respectively. Read the documentation about volumes for more information. Essentially, -v /path/to/matrix/on/host:/input_matrix with -m /input_matrix is what you want, where before the : is on the host filesystem while after the : is what the docker program sees. Then you can write the output in the same way: -v /path/to/output/on/host:/output will write the output to the folder before the :.

To build the too-many-cells image yourself if you want:

git clone https://github.com/GregorySchwartz/too-many-cells.git
cd too-many-cells
docker build -t too-many-cells -f ./Dockerfile .

macOS

We recommend using docker on macOS. If you need to build too-many-cells, you should get the above dependencies. For some dependencies, you can use brewer, then install too-many-cells (in the cloned folder, don't forget to install the R dependencies above):

brew cask install xquartz
brew install glib cairo gtk gettext fontconfig freetype

brew tap brewsci/bio
brew tap brewsci/science
brew install r zeromq graphviz pkg-config gsl libffi gobject-introspection gtk+ gtk+3

# Needed so pkg-config and libraries can be found.
# For the second path, use the ouput of "brew info libffi".
export PKG_CONFIG_PATH=/usr/local/lib/pkgconfig:/usr/local/opt/libffi/lib/pkgconfig

# Tell gtk that it's quartz
stack install --flag gtk:have-quartz-gtk

Troubleshooting

I am getting errors like AesonException "Error in $.packages.cassava.constraints.flags... when running stack commands

Try upgrading stack with stack upgrade. The new installation will be in ~/.local/bin, so use that binary.

I use conda or custom ld library locations and I cannot install too-many-cells or run into weird R errors

stack and too-many-cells assume system libraries and programs. To solve this issue, first install the dependencies above at the system level, including system R. Then to every stack and too-many-cells command, prepend PATH="$HOME/.local/bin:/usr/bin:$PATH" to all commands. For instance:

  • PATH="$HOME/.local/bin:/usr/bin:$PATH" stack install
  • PATH="$HOME/.local/bin:/usr/bin:$PATH" too-many-cells make-tree -h

If your shared libraries are abnormal and use libR.so from non-system locations, be sure to also have LD_LIBRARY_PATH=/usr/lib/:$LD_LIBRARY_PATH when installing.

I am still having issues with installation

Open an issue! While working on the issue, try out the docker for too-many-cells, it requires no installation at all (other than docker).

Included projects

This project is a collection of libraries and programs written specifically for too-many-cells:

birch-beer
Generate a tree for displaying a hierarchy of groups with colors, scaling, and more.
modularity
Find the modularity of a network.
spectral-clustering
Library for spectral clustering.
hierarchical-spectral-clustering
Hierarchical spectral clustering of a graph.
differential
Finds out whether an entity comes from different distributions (statuses).

Usage

too-many-cells has several entry points depending on the desired analysis.

Argument Analysis
make-tree Generate the tree from single cell data with various measurement outputs and visualize tree
interactive Interactive visuzalization of the tree, very slow
differential Find differentially expressed genes between two nodes
diversity Conduct diversity analyses of multiple cell populations
paths The binary tree equivalent of the so called "pseudotime", or 1D dimensionality reduction

The main workflow is to first generate and plot the population tree using too-many-cells make-tree, then use the rest of the entry points as needed.

At any point, use -h to see the help of each entry point.

Also, check out tooManyCellsR for an R wrapper!

make-tree

too-many-cells make-tree generates a binary tree using hierarchical spectral clustering. We start with all cells in a single node. Spectral clustering partitions the cells into two groups. We assess the clustering using Newman-Girvan modularity: if \(Q > 0\) then we recursively continue with hierarchical spectral clustering. If not, then there is only a single community and we do not partition – the resulting node is a leaf and is considered the finest-grain cluster.

The most important argument is the --prior argument. Making the tree may take some time, so if the tree was already generated and other analysis or visualizations need to be run on the tree, point the --prior argument to the output folder from a previous run of too-many-cells. If you do not use --prior, the entire tree will be recalculated even if you just wanted to change the visualization!

The main input is the --matrix-path argument. When a directory is supplied, too-many-cells interprets the folder to have matrix.mtx, genes.tsv, and barcodes.tsv files (cellranger outputs, see cellranger for specifics). If a file is supplied instead of a directory, we assume a csv file containing gene row names and cell column names. This argument can be called multiple times to combine multiple single cell matrices: --matrix-path input1 --matrix-path input2.

The second most important argument is --labels-file. Supply with a csv with a format and header of "item,label" to provide colorings and statistics of the relationships between labels. Here the "item" column contains the name of each cell (barcode) and the label is any property of the cell (the tissue of origin, hour in a time course, celltype, etc.).

To see the full list of options, use too-many-cells -h and -h for each entry point (i.e. too-many-cells make-tree -h).

Output

too-many-cells make-tree generates several files in the output folder. Below is a short description of each file.

File Description
clumpiness.csv When labels are provided, uses the clumpiness measure to determine the level of aggregation between each label within the tree.
clumpiness.pdf When labels are provided, a figure of the clumpiness between labels.
cluster_diversity.csv When labels are provided, the diversity, or "effective number of labels", of each cluster.
cluster_info.csv Various bits of information for each cluster and the path leading up to each cluster, from that cluster to the root. For instance, the size column has cluster_size/parent_size/parent_parent_size/.../root_size
cluster_list.json The json file containing a list of clusterings.
cluster_tree.json The json file containing the output tree in a recursive format.
dendrogram.svg The visualization of the tree. There are many possible options for this visualization included. Can rename to choose between PNG, PS, PDF, and SVG using --dendrogram-output.
graph.dot A dot file of the tree, with less information than the tree in cluster_results.json.
node_info.csv Various information of each node in the tree.
projection.pdf When --projection is supplied with a file of the format "barcode,x,y", provides a plot of each cell at the specified x and y coordinates (for instance, when looking at t-SNE plots with the same labelings as the dendrogram here).

Outline with options

The basic outline of the default pre-processing pipeline with some relevant options is as follows (there are many additional options including cell whitelists and PCA that can be seen using too-many-cells make-tree -h):

  1. Read matrix.
  2. Remove cells with less than 250 counts (--filter-thresholds, --no-filter).
  3. Remove genes with less than 1 count (--filter-thresholds, --no-filter).
  4. Term frequency-inverse document frequency normalization (--normalization).
  5. Finish.

Example

  • Setup

    We start with our input matrix. Here,

    ls ./input
    
    barcodes.tsv  genes.tsv  matrix.mtx
    

    Note that the input can be a directory (with the cellranger matrix format above) or a file (a csv file). You can also point to a cellranger >= 3.0 folder which has matrix.mtx.gz, features.tsv.gz, and barcodes.tsv.gz files instead. You don't need to use scRNA-seq data! You can use any data that has observations (cells) and features (genes), as long as you agree that the observations are related by their feature abundances. If you do upstream batch effect correction, PCA, normalization, or anything else, be sure to use --no-filter --normalization NoneNorm (and --shift-positive for PCA) to avoid wrong filters and scalings! If using dimensionality reduction such as PCA and t-SNE, we highly recommend generating your own similarity matrix for use with our cluster-tree program and plot with birch-beer, as we emphasize a feature matrix in too-many-cells and dimensionality reduction algorithms transform counts (our input which works with cosine similarity) into more nebulous information (which may not work with cosine similarity). cluster-tree, however, can be used with adjacency and similarity matrices. As for formats, the matrix market format contains three files like so:

    The matrix.mtx file is in matrix market format.

    %%MatrixMarket matrix coordinate integer general
    %
    23433 1981 4255069
    4 1 1
    5 1 1
    11 1 2
    23 1 2
    25 1 2
    40 1 2
    48 1 1
    ...
    

    The genes.tsv file (or features.tsv.gz) contains the features of each cell and corresponds to the rows of matrix.mtx. Here, both columns were the same gene symbols, but you can have Ensembl as the first column and gene symbol as the second, etc. The columns and column orders don't matter, but make sure all matrices have the same format and specify the symbols you want to use (for overlaying gene expression, differential expression, etc.) with --feature-column COLUMN. So to use the second column for gene expression, you would use --feature-column 2.

    Xkr4	Xkr4
    Rp1	Rp1
    Sox17	Sox17
    Mrpl15	Mrpl15
    Lypla1	Lypla1
    Tcea1	Tcea1
    Rgs20	Rgs20
    Atp6v1h	Atp6v1h
    Oprk1	Oprk1
    Npbwr1	Npbwr1
    ...
    

    The barcodes.tsv file contains the ids of each cell or observation and corresponds to the columns of matrix.mtx.

    AAACCTGCAGTAACGG-1
    AAACGGGAGAAGAAGC-1
    AAACGGGAGACCGGAT-1
    AAACGGGAGCGCTCCA-1
    AAACGGGAGGACGAAA-1
    AAACGGGAGGTACTCT-1
    AAACGGGAGGTGCTTT-1
    AAACGGGAGTCGAGTG-1
    AAACGGGCATGGTCAT-1
    AAAGATGAGCTTCGCG-1
    ...
    

    For a csv file, the format is dense (observation columns (cells), feature rows (genes)):

    "","A22.D042044.3_9_M.1.1","C5.D042044.3_9_M.1.1","D10.D042044.3_9_M.1.1","E13.D042044.3_9_M.1.1","F19.D042044.3_9_M.1.1","H2.D042044.3_9_M.1.1","I9.D042044.3_9_M.1.1",...
    "0610005C13Rik",0,0,0,0,0,0,0,...
    "0610007C21Rik",0,112,185,54,0,96,42,...
    "0610007L01Rik",0,0,0,0,0,153,170,...
    "0610007N19Rik",0,0,0,0,0,0,0,...
    "0610007P08Rik",0,0,0,0,0,19,0,...
    "0610007P14Rik",0,58,0,0,255,60,0,...
    "0610007P22Rik",0,0,0,0,0,65,0,...
    "0610008F07Rik",0,0,0,0,0,0,0,...
    "0610009B14Rik",0,0,0,0,0,0,0,...
    ...
    

    We also know where each cell came from, so we mark that down as well in a labels.csv file.

    item,label
    AAACCTGCAGTAACGG-1,Marrow
    AAACGGGAGACCGGAT-1,Marrow
    AAACGGGAGCGCTCCA-1,Marrow
    AAACGGGAGGACGAAA-1,Marrow
    AAACGGGAGGTACTCT-1,Marrow
    ...
    

    This can be easily accomplished with sed:

    cat barcodes.tsv | sed "s/-1/-1,Marrow/" | s/-2/etc... > labels.csv
    

    For cellranger, note that the -1, -2, etc. postfixes denote the first, second, etc. label in the aggregation csv file used as input for cellranger aggr.

  • Default run

    We can now run the too-many-cells algorithm on our data. The resulting cells with assigned clusters will be printed to stdout (don't forget to use --no-filter and --normalization NoneNorm on preprocessed data, as stated here).

    too-many-cells make-tree \
        --matrix-path input \
        --labels-file labels.csv \
        --draw-collection "PieRing" \
        --output out \
        > clusters.csv
    

    complete_default_tree.png

  • Pruning tree

    Large cell populations can result in a very large tree. What if we only want to see larger subpopulations rather than the large (inner nodes) and small (leaves)? We can use the --min-size 100 argument to set the minimum size of a leaf to 100 in this case. Alternatively, we can specify --smart-cutoff 4 in addition to --min-size 1 to set the minimum size of a node to \(4 * \text{median absolute deviation (MAD)}\) of the nodes in the original tree. Varying the number of MADs varies the number of leaves in the tree. --smart-cutoff should be used in addition to --min-size, max-proportion, or min-distance to decide which cutoff variable to use. The value supplied to the cutoff variable is ignored when --smart-cutoff is specified. We'll prune the tree for better visibility in this document.

    Note: the pruning arguments change the tree file, not just the plot, so be sure to output into a different directory.

    Also, we do not need to recalculate the entire tree! We can just supply the previous results using --prior (we can also remove --matrix-path with --prior to speed things up, but miss out on some features if needed):

    too-many-cells make-tree \
        --prior out \
        --labels-file labels.csv \
        --smart-cutoff 4 \
        --min-size 1 \
        --draw-collection "PieRing" \
        --output out_pruned \
        > clusters_pruned.csv
    

    pruned_tree.png

  • Pie charts

    What if we want pie charts instead of showing each individual cell (the default)?

    too-many-cells make-tree \
        --prior out \
        --labels-file labels.csv \
        --smart-cutoff 4 \
        --min-size 1 \
        --draw-collection "PieChart" \
        --output out_pruned \
        > clusters_pruned.csv
    

    piechart_pruned_tree.png

  • Node numbering

    Now that we see the relationships between clusters and nodes in the dendrogram, how can we go back to the data – which nodes represent which node IDs in the data?

    too-many-cells make-tree \
        --prior out \
        --labels-file labels.csv \
        --smart-cutoff 4 \
        --min-size 1 \
        --draw-collection "PieChart" \
        --draw-node-number \
        --output out_pruned \
        > clusters_pruned.csv
    

    numbered_pruned_tree.png

  • Branch width

    We can also change the width of the nodes and branches, for instance if we want thinner branches:

    too-many-cells make-tree \
        --prior out \
        --labels-file labels.csv \
        --smart-cutoff 4 \
        --min-size 1 \
        --draw-collection "PieChart" \
        --draw-max-node-size 40 \
        --output out_pruned \
        > clusters_pruned.csv
    

    thin_pruned_tree.png

  • No scaling

    We can remove all scaling for a normal tree and still control the branch widths:

    too-many-cells make-tree \
        --prior out \
        --labels-file labels.csv \
        --smart-cutoff 4 \
        --min-size 1 \
        --draw-collection "PieChart" \
        --draw-max-node-size 40 \
        --draw-no-scale-nodes \
        --output out_pruned \
        > clusters_pruned.csv
    

    no_scaling_pruned_tree.png

    How strong is each split? We can tell by drawing the modularity of the children on top of each node:

    too-many-cells make-tree \
        --prior out \
        --labels-file labels.csv \
        --smart-cutoff 4 \
        --min-size 1 \
        --draw-collection "PieChart" \
        --draw-mark "MarkModularity" \
        --output out_pruned \
        > clusters_pruned.csv
    

    modularity_pruned_tree.png

  • Gene expression

    What if we want to draw the gene expression onto the tree in another folder (requires --matrix-path, may take some time depending on matrix size. Defaults to all black if the feature name is not present in the matrix, so check the first column of the feature file)? Note: the feature names are from the genes.tsv or features.tsv.gz file. Usually, cellranger has Ensembl identifiers as the first column and gene symbol as the second column, so if you want to specify gene symbol, use --feature-column 2 (1 is default).

    too-many-cells make-tree \
        --prior out \
        --matrix-path input \
        --labels-file labels.csv \
        --smart-cutoff 4 \
        --min-size 1 \
        --feature-column 2 \
        --draw-leaf "DrawItem (DrawContinuous [\"Cd4\"])" \
        --output out_gene_expression \
        > clusters_pruned.csv
    

    cd4_dendrogram.png

    Notice that Cd4 is within a list ([]), so multiple features can be listed and the average of those values for each cell will be used. While this representation shows the expression of Cd4 in each cell and blends those levels together, due to the sparsity of single cell data these cells and their respective subtrees may be hard to see without additional processing. Let's scale the saturation to more clearly see sections of the tree with our desired expression (when choosing other high and low colors with --draw-colors, scaling the saturation will only affect non-grayscale colors).

    too-many-cells make-tree \
        --prior out \
        --matrix-path input \
        --labels-file labels.csv \
        --smart-cutoff 4 \
        --min-size 1 \
        --feature-column 2 \
        --draw-leaf "DrawItem (DrawContinuous [\"Cd4\"])" \
        --draw-scale-saturation 10
        --output out_gene_expression \
        > clusters_pruned.csv
    

    cd4_saturated_10_dendrogram.png

    There, much better! Now it's clearly enriched in the subtree containing the thymus, where we would expect many T cells to be. While this tree makes the expression a bit more visible, there is another tactic we can use. Instead of the continuous color spectrum of expression values, we can have a binary "high" and "low" expression. Here, we'll continue to have the red and gray colors represent high and low expressions respectively using the --draw-colors argument. Note that this binary expression technique can be used for multiple features, hence it's a list of features with cutoffs so you can be high in a gene and low in another gene, etc. for all possible combinations.

    too-many-cells make-tree \
        --prior out \
        --matrix-path input \
        --labels-file labels.csv \
        --smart-cutoff 4 \
        --min-size 1 \
        --feature-column 2 \
        --draw-leaf "DrawItem (DrawThresholdContinuous [(\"Cd4\", 0), (\"Cd8a\", 0)])" \
        --draw-colors "[\"#e41a1c\", \"#377eb8\", \"#4daf4a\", \"#eaeaea\"]" \
        --draw-scale-saturation 10 \
        --output out_gene_expression \
        > clusters_pruned.csv
    

    cd4_cd8_sat_10_dendrogram.png

    Now we can see the expression of both Cd4 and Cd8a at the same time!

  • Diversity

    We can also see an overview of the diversity of cell labels within each subtree and leaves.

    too-many-cells make-tree \
        --prior out \
        --matrix-path input \
        --labels-file labels.csv \
        --smart-cutoff 4 \
        --min-size 1 \
        --draw-leaf "DrawItem DrawDiversity" \
        --output out_diversity \
        > clusters_pruned.csv
    

    diversity_pruned_tree.png

    Here, the deeper the red, the more diverse (a larger "effective number of cell states") the cell labels in that group are. Note that the inner nodes are colored relative to themselves, while the leaves are colored relative to all leaves, so there are two different scales.

interactive

The interactive entry point has a basic GUI interface for quick plotting with a few features. We recommend limited use of this feature, however, as it can be quite slow at this stage, has fewer customizations, and requires specific dependencies.

too-many-cells interactive \
    --prior out \
    --labels-file labels.csv

differential

A main use of single cell clustering is to find differential genes between multiple groups of cells. The differential aids in this endeavor by allowing comparisons with edgeR. Let's find the differential genes between the liver group and all other cells. Consider our pruned tree from earlier:

piechart_pruned_tree.png

We can see the id of each group with --draw-node-number.

numbered_pruned_tree.png

We need to define two groups to compare. Well, it looks like node 98 defines the liver cluster. Then, since we don't want 98 to be in the other group, we say that all other cells are within nodes 89 and 1. As a result, we end up with a tuple containing two lists: ([89, 1], [98]). Then our differential genes for (liver / others) can be found with differential (sent to stdout):

too-many-cells differential \
    --matrix-path input \
    -n "([89, 1], [98])" \
    > differential.csv

If we wanted to make the same comparison, but compare the liver subtree with liver cells from all other subtrees, we can use the --labels argument:

too-many-cells differential \
    --matrix-path input \
    --labels-file labels.csv \
    -n "([89, 1], [98])" \
    --labels "([\"Liver\"], [\"Liver\"])" \
    > differential_liver.csv

We can also look at the distribution of abundance for individual genes using the --genes and --plot-output arguments.

Furthermore, we can compare each node to all other cells by specifying no nodes at all. The output file will contain the top --top-n genes for each node. We recommend using multiple OS threads here to speed up the process using +RTS -N${NUMOSTHREADS} (no number to use all cores). The following example will compare all nodes to all other cells using 8 OS threads:

too-many-cells differential \
    --matrix-path input \
    -n "([], [])" \
    --normalization "UQNorm" \
    +RTS -N8

diversity

Diversity is the measure of the "effective number of entities within a system", originating from ecology (See Jost: Entropy and Diversity). Here, each cell is an organism and each cell label or cluster is a species, depending on the question. In ecology, the diversity index measures the effective number of species within a population such that the minimum is a diversity of 1 for a single dominant species up to maximum of the total number of species (evenly abundant). If our species is a cluster, then here the diversity is the effective number of cell states within a population (for labels, make-tree generates these results automatically in "diversity" columns). Say we have two populations and we generated the trees using make-tree into two different output folders, out1 and out2. We can find the diversity of each population using the diversity entry point.

too-many-cells diversity\
    --priors out1 \
    --priors out2 \
    -o out_diversity_stats

We can then find a simple plot of diversity in diversity_output. In addition, we also provide rarefaction curves for comparing the number of different cell states at each subsampling useful for comparing the number of cell states where the population sizes differ.

paths

"Pseudotime" refers to the one dimensional relationship between cells, useful for looking at the ordering of cell states or labels. The implementation of pseudotime in a too-many-cells point-of-view is by finding the distance between all cells and the cells found in the longest path from the root in the tree. Then each cell has a distance from the "start" and thus we plot those distances.

too-many-cells paths\
    --prior out \
    --labels-file labels.csv \
    --bandwidth 3 \
    -o out_paths

Advanced documentation

Each entry point has its own documentation accessible with -h, such as too-many-cells make-tree -h:

too-many-cells -h
too-many-cells, Gregory W. Schwartz. Clusters and analyzes single cell data.

Usage: too-many-cells (make-tree | interactive | differential | diversity |
                      paths)

Available options:
  -h,--help                Show this help text

Available commands:
  make-tree
  interactive
  differential
  diversity
  paths

Demo

Check out an instructional example of using too-many-cells here when finished looking at the brief feature overview.

Author: Gregory W. Schwartz

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