Demuxlet¶
This workflow runs demuxlet to deconvolute sample identity when multiple samples are pooled by barcoded single-cell sequencing.
Align your single-cell sequencing data (for example using the cellranger or drop_seq workflows).
Create a sample sheet.
Please note that the columns in the tab separated file must be in the order shown below and does not contain a header line.
Column Description Name Sample name. BAM Location of the BAM file in the cloud (gs:// URL). Barcodes Location of the valid cellular barcodes file in the cloud (gs:// URL). VCF Location of the VCF file to use for this sample in the cloud (gs:// URL). Example:
sample-1,gs://fc-e0000000/sample-1/out/possorted_genome_bam.bam,gs://fc-e0000000/sample-1/out/filtered_feature_bc_matrix/barcodes.tsv.gz,gs://fc-e0000000/sample-1.vcf sample-2,gs://fc-e0000000/sample-2/out/possorted_genome_bam.bam,gs://fc-e0000000/sample-2/out/filtered_feature_bc_matrix/barcodes.tsv.gz,gs://fc-e0000000/sample-2.vcf
Upload your sample sheet to the workspace bucket.
Example:
gsutil cp /foo/bar/projects/sample_sheet.tsv gs://fc-e0000000/
Import demuxlet workflow to your workspace.
See the Terra documentation for adding a workflow. The workflow is under
Broad Methods Repository
with the name “cumulus/demuxlet”.Next, in the workflow page, click the
Export to Workspace...
button, and select the workspace you want to export to in the drop-down menu.In your workspace, open
demuxlet
inWORKFLOWS
tab. SelectProcess single workflow from files
as belowand click the
Save
button.
Inputs¶
Please see the description of important inputs below.
Column | Description |
---|---|
tsv_file | Four column tab-separated file without a header with name, coordinate sorted bam, barcodes, and vcf |
min_MQ | Minimum mapping quality to consider (default 20) |
alpha | Grid of alpha to search for (default [0.1, 0.2, 0.3, 0.4, 0.5]). |
min_TD | Minimum distance to the tail (default 0) |
tag_group | Tag representing readgroup or cell barcodes, in the case to partition the BAM file into multiple groups (default “CB”) |
tag_UMI | Tag representing UMIs (default “UB”“) |
field | FORMAT field to extract the genotype, likelihood, or posterior from (default “GT”) |
geno_error | Offset of genotype error rate (default 0.1) |
Outputs¶
The demuxlet output file contains the best guess of the sample identity, with detailed statistics to reach to the best guess.