Samples Table

Field	Used for Multiome	Used for RNA-seq	Used for ATAC-seq	Required
sample	yes	yes	yes	yes
replicate	optional	optional	optional	optional
genome	yes	no	yes	yes, ATAC/Multiome
HDF5_Multiple_Assay	yes	yes	yes	See notes below
RDS_Multiple_Assays	yes	yes	yes	See notes below
Gene.Expression	yes	yes	no	See notes below
Peaks	yes	no	yes	See notes below
TF	yes	no	yes	See notes below
fragments	yes	no	yes	yes, ATAC/Multiome
singlecell	yes	no	yes	no
meta_batch	optional	optional	optional	optional
meta_geno	optional	optional	optional	optional

Note

• Suppored input file formats include HDF5 (10X Genomics compatible), MEX directory, or RDS.

• For input from non-10X Genomics pipelines, place the barcodes.tsv.gz, features.tsv.gz, and matrix.mtx.gz in the MEX directory.

• For details about 10X Genomics’ HDF5 format, refer to the HDF5 Feature-Barcode Matrix Format.

Field descriptions

sample

string. Defines labels for each sample. Values in sample column must be unique, unless analyzing technical replicates. If specifying technical replicates, sample label must be the same for all rows containing techical replicates.

replicate

string. Optional. Defines labels for each technical replicate of a sample. Values in replicate column must be unique per sample. If not specified technical replicates, leave empty. If specified technical replicates: sample must be the same for all rows containing techical replicates.

Note

Refer to the Replicates section for the definition of biological and technical replicates.

genome

string, default hg38. Defines labels for genome build per sample. If samples include “Peaks” or “Transcription Factor” (TF) matrices, as with ATAC and Multiome products, all samples must have genome. Otherwise, genome labels can differ across samples. In the current version of the multiome-wf, mm10 (mouse) and hg38 (human) are available.

HDF5_Multiple_Assays

Path to HDF5 file containing multiple feature-by-barcode matrices. If a sample’s input contains multiple matrices, list the sample path in either HDF5_Multiple_Assays or RDS_Multiple_Assays columns of samples.tsv. Paths specified in HDF5_Multiple_Assays should point to a 10X Genomics compatible HDF5 file.

Note

• If the specified input file contains feature-by-barcode matrices f or 5’ or 3’ Gene Expression counts matrix, or ATAC/Multiome counts matrices including Peaks or Transcription Factors, do not duplicate paths in Gene.Expression, Peaks or TF columns, respectively.

• Specifying paths in this column is useful if you are analyzing 10X Genomics kits containing multiple assays, such as Gene Expression + CRISPR barcodes (Perturb-Seq), 3’ Gene Expression + Protein barcodes (CITE-Seq), Multiome (Gene Expression + ATAC), etc.

• Specifying paths in this column can also be useful if you are analyzing data derived from non-10X Genomics platforms, which may need reformatting prior to running the workflow.

RDS_Multiple_Assays

Path to RDS file containing multiple feature-by-barcode matrices saved in a named list of dgCMatrices. The saved data format is identical to that of HDF5_Multiple_Assays (Refer to the notes under HDF5_Multiple_Assays).

Gene.Expression

Path to a single feature-by-barcode matrix containing 5’ or 3’ Gene Expression counts. Path can point to either 1) a file in HDF or RDS format, or 2) the parent directory containing a 10X compatible MEX sparse matrix and associated barcodes and features files.

Note

• Specifying paths in this column is useful if you are analyzing 10X Genomics single-assay transcriptomics kits.

• If analyzing single cell transcriptomics data from a non-10X Genomics platform, reformatting the gene-by-barcode count matrix is likely needed. In such a case, the reformatted counts matrix can be save in HDF5 or RDS format and specified in the Gene.Expression column of samples.tsv.

• If the specified matrix has any additional features, for example 3’ Gene Expression + CRISPR barcodes (Perturb-Seq), 3’ Gene Expression + Protein barcodes (CITE-Seq), use either HDF5_Multiple_Assays or RDS_Multiple_Assays columns of samples.tsv.

Peaks

Path to a single feature-by-barcode matrix containing ATAC peaks counts. Path can point to either 1) a file in HDF or RDS format, or 2) the parent directory containing a 10X compatible MEX sparse matrix and associated barcodes and features files.

Note

• Specifying paths in this column is useful if you are analyzing 10X Genomics single-assay chromatin accessibility kits.

• If analyzing single cell ATAC data from a non-10X platform, reformatting the gene-by-barcode count matrix is likely needed. In such a case, the reformatted counts matrix can be save in HDF5 or RDS format and specified in the Peaks column of samples.tsv.

• If the specified matrix has any additional features, for example Multiome (Gene Expression + ATAC), use either HDF5_Multiple_Assays or RDS_Multiple_Assays columns of samples.tsv.

TF

Path to a single feature-by-barcode matrix encoding Transcription Factor counts. Path can point to either 1) a file in HDF or RDS format, or 2) the parent directory containing a 10X compatible MEX sparse matrix and associated barcodes and features files. Refer to the notes under Peaks.

fragments

Path to the fragments.tsv.gz file containing a table of tagmentation loci, each with coordinates and de-duplicated counts, created from 10X Genomics ATAC and Multiome kits.

singlecell

Path to the singlecell.csv file created from 10X Genomics ATAC and Multiome kits.

meta_*

string. Optional. Define columns for metadata labels. Columns beginning with meta_ are placeholders for user-specified metadata. Users can rename these columns.

Note

• In samples.tsv, the following column names are considered immutable: sample, replicate, genome, HDF5_Multiple_Assays, RDS_Multiple_Assays, Gene.Expression, Peaks, TF, fragments, singlecell.

• Any additional columns present in samples.tsv will be considered metadata columns. Metadata columns can have any unique label, not just the meta_* used in the example samples table.

Examples

Multiome

A basic examples of a samples.tsv file for 10X Genomics Multiome analysis is below:

sample	replicate	genome	HDF5_Multiple_Assays	RDS_Multiple_Assays	Gene.Expression	Peaks	TF	fragments	singlecell	meta_batch	meta_geno
rep1_wt		mm10	rep1_wt/outs/filtered_feature_bc_matrix.h5					rep1_wt/outs/atac_fragments.tsv.gz		batch1	wt
rep1_homo		mm10	rep_homo/outs/filtered_feature_bc_matrix.h5					rep1_homo/outs/atac_fragments.tsv.gz		batch1	ko

RNA-seq

A basic examples of a samples.tsv file for 10X Genomics Single Cell 3’ Gene Expression analysis is below:

sample	replicate	genome	HDF5_Multiple_Assays	RDS_Multiple_Assays	Gene.Expression	Peaks	TF	fragments	singlecell	meta_batch	meta_tissue
CTX		mm10			wt/outs/filtered_feature_bc_matrix.h5					batch1	CTX
MGE		mm10			ko/outs/filtered_feature_bc_matrix.h5					batch1	MGE

ATAC-seq

A basic examples of a samples.tsv file for 10X Genomics Single Cell ATAC analysis is below:

sample	replicate	genome	HDF5_Multiple_Assays	RDS_Multiple_Assays	Gene.Expression	Peaks	TF	fragments	singlecell	meta_batch	meta_tissue
CTX		mm10				CTX_rerun/outs/filtered_peak_bc_matrix.h5	CTX_rerun/outs/filtered_tf_bc_matrix.h5	CTX_rerun/outs/fragments.tsv.gz	CTX_rerun/outs/summary.csv	batch1	CTX
MGE		mm10				MGE_rerun/outs/filtered_peak_bc_matrix.h5	MGE_rerun/outs/filtered_tf_bc_matrix.h5	MGE_rerun/outs/fragments.tsv.gz	MGE_rerun/outs/summary.csv	batch1	MGE

See Overview of workflows for more detailed examples of config files.