Options and Arguments#
It is important to tweak parameter and arguments. Usually the defaults are fine but changing resolution and MT trimming percentage can affect the results.
Cellsnake generates different directories for these main parameters so you can check their effects on the results.
If you decide a value, you can delete the unnecessary directories and keep the others for integration or later use.
For example, if you run cellsnake with the following parameters:
#data here is a directory with your input files, there can be multiple directories, symlinks are allowed.
#this command will generate results under folder: results/sample_name/percent_mt~5/resolution~0.6/.
cellsnake minimal data --percent_mt 5 --resolution 0.6
Modes#
Cellsnake has 5 modes: minimal, standard, advanced, integration and clustree.
Mode |
Outputs |
|---|---|
minimal |
Dimension reduction plots, QC metrics, technical plots (MT, counts, gene, feature), clustree plot |
standard |
All minimal outputs and CellTypist, singleR annotations, enrichment analyses tables, trajectory plots, summarized marker plots |
advanced |
All standard outputs and CellChat results, detailed top markers per cluster plots |
clustree |
Clustree plot and basic QC plots |
integrate |
A single integrated object for integrated analysis |
Full parameter list#
Usage:
cellsnake <command> <INPUT> [options] [--unlock|--remove] [--dry]
cellsnake integrated <command> <INPUT> [options] [--unlock|--remove] [--dry]
cellsnake --generate-template
cellsnake --install-packages
cellsnake (-h | --help)
cellsnake --version
commands:
minimal Run cellsnake with minimal workflow.
standard Run cellsnake with standard workflow.
advanced Run cellsnake with advanced workflow.
clustree Run cellsnake with clustree workflow.
integrate Run cellsnake to integrate samples under analyses folder.
This option expects you have already finished processing multiple samples.
main arguments:
INPUT Input directory or a file to process (if a directory given, batch mode is ON).
--configfile <text> Config file name in YAML format, for example, "config.yaml". No default but can be created with --generate-template.
--metadata <text> Metadata file name in CSV, TSV or Excel format, for example, "metadata.csv", header required, first column sample name. No default but can be created with --generate-template.
--metadata_column <text> Metadata column for differential expression analysis [default: condition].
other arguments:
--gene <gene or filename> Create publication ready plots for a gene or a list of genes from a text file.
main options:
--percent_mt <double> Maximum mitochondrial gene percentage cutoff,
for example, 5 or 10, write "auto" for auto detection [default: 10].
--resolution <double> Resolution for cluster detection, write "auto" for auto detection [default: 0.8].
other options:
--doublet_filter <bool> [default: True] #this may fail on some samples
--percent_rp <double> [default: 0] #Ribosomal genes minimum percentage (0-100), default no filtering
--min_cells <integer> [default: 3] #seurat default, recommended
--min_features <integer> [default: 200] #seurat default, recommended, nFeature_RNA
--max_features <integer> [default: Inf] #seurat default, nFeature_RNA, 5000 can be a good cutoff
--min_molecules <integer> [default: 0] #seurat default, nCount_RNA, min_features usually handles this so keep it 0
--max_molecules <integer> [default: Inf] #seurat default, nCount_RNA, to filter potential doublets, doublet filtering is already default, so keep this Inf
--highly_variable_features <integer> [default: 2000] #seurat defaults, recommended
--variable_selection_method <text> [default: vst] #seurat defaults, recommended
--normalization_method <text> [default: LogNormalize]
--scale_factor <integer> [default: 10000]
--logfc_threshold <double> [default: 0.25]
--test_use <text> [default: wilcox]
--mapping <text> [default: org.Hs.eg.db] #you may install others from Bioconductor, this is for human
--organism <text> [default: hsa] #alternatives https://www.genome.jp/kegg/catalog/org_list.html
--species <text> [default: human] for cellchat, #only human or mouse is accepted
plotting parameters:
--min_percentage_to_plot <double> [default: 2] #only show clusters more than % of cells on the legend
--show_labels <bool> [default: True] #
--marker_plots_per_cluster_n <integer> [default: 20] #plot summary marker plots for top markers
--umap_markers_plot <bool> [default: True]
--tsne_markers_plot <bool> [default: False]
annotation options:
--singler_ref <text> [default: BlueprintEncodeData] # https://bioconductor.org/packages/release/data/experiment/vignettes/celldex/inst/doc/userguide.html#1_Overview
--celltypist_model <text> [default: Immune_All_Low.pkl] #refer to Celltypist for another model
microbiome options:
--kraken_db_folder <text> No default, you need to provide a folder with kraken2 database
--taxa <text> [default: genus] # available options "domain", "kingdom", "phylum", "class", "order", "family", "genus", "species"
--microbiome_min_cells <integer> [default: 1]
--microbiome_min_features <integer> [default: 3]
--confidence <double> [default: 0.05] #see kraken2 manual
--min_hit_groups <integer> [default: 4] #see kraken2 manual
integration options:
--dims <integer> [default: 30] #refer to Seurat for more details
--reduction <text> [default: cca] #refer to Seurat for more details
others:
--generate-template Generate config file template and metadata template in the current directory.
--install-packages Install, reinstall or check required R packages.
-j <integer>, --jobs <integer> Total CPUs. [default: 2]
-u, --unlock Rescue stalled jobs (Try this if the previous job ended prematurely or currently failing).
-r, --remove Delete all output files (this won't affect input files).
-d, --dry Dry run, nothing will be generated.
-h, --help Show this screen.
--version Show version.