Cell Ranger6.0, printed on 11/21/2024
Cell Ranger performs analysis of single-cell Targeted Gene Expression data with the cellranger count pipeline. Cell Ranger provides the same interface and features for both Targeted and Whole Transcriptome Analysis (WTA) Gene Expression data, as described in Single-Library Analysis with Cell Ranger.
To enable Targeted Gene Expression analysis in cellranger count, specify the relevant Target Panel CSV file using the --target-panel flag:
$ cd /home/jdoe/runs $ cellranger count --id=sample345 \ --target-panel=/opt/cellranger-5.0.0/target_panels/immunology_v1.0_GRCh38-2020-A.target_panel.csv \ --transcriptome=/opt/refdata-gex-GRCh38-2020-A \ --fastqs=/home/jdoe/runs/HAWT7ADXX/outs/fastq_path \ --sample=mysample \ --expect-cells=1000 \ --localcores=8 \ --localmem=64
Target Panel CSV files corresponding to predesigned gene panels can be found within the target_panels directory in the Cell Ranger package. For example, if you installed Cell Ranger to /opt/cellranger-5.0.0, then the target panel CSVs will be contained in /opt/cellranger-5.0.0/target_panels. These CSV files can also be downloaded on the Panel Selection page, along with more detailed information about each panel.
Detailed documentation on this file format can be found here, along with descriptions of all other downloads provided for target gene panels.
The output from a Targeted Gene Expression analysis is generally very similar to output from non-targeted analysis—see the table below for a comprehensive list of differences. The primary changes are the addition of targeting metrics, and that all secondary analysis for targeted data is performed after removing non-targeted genes from the count matrix. Likewise, the filtered feature-barcode matrix files and Loupe Browser file do not include non-targeted gene counts. The raw feature-barcode matrices and molecule info H5 file still provide information about all genes.
| File Name | Description | Differences for Targeted Gene Expression |
|---|---|---|
web_summary.html | Run summary metrics and charts in HTML format | Contains additional Targeted Gene Expression metrics |
metrics_summary.csv | Run summary metrics in CSV format | Contains additional Targeted Gene Expression metrics |
possorted_genome_bam.bam | Reads aligned to the genome and transcriptome annotated with barcode information | The xf tag now indicates whether a read was filtered by targeted UMI filtering |
possorted_genome_bam.bam.bai | Index for possorted_genome_bam.bam | |
filtered_feature_bc_matrix | Filtered feature-barcode matrices containing only cellular barcodes in MEX format | Contains only targeted genes |
filtered_feature_bc_matrix_h5.h5 | Filtered feature-barcode matrices containing only cellular barcodes in HDF5 format | Contains only targeted genes* |
raw_feature_bc_matrices | Unfiltered feature-barcode matrices containing all barcodes in MEX format | |
raw_feature_bc_matrix_h5.h5 | Unfiltered feature-barcode matrices containing all barcodes in HDF5 format | Generally unchanged* |
analysis | Secondary analysis data including dimensionality reduction, cell clustering, and differential expression | Secondary analysis performed only on the targeted genes |
molecule_info.h5 | Molecule-level information used by cellranger aggr to aggregate samples into larger datasets. | Generally unchanged* |
cloupe.cloupe | Loupe Browser visualization and analysis file | Contains only targeted genes |
target_panel.csv | Target Panel CSV file | Only present for Targeted Gene Expression data |
feature_reference.csv | (Feature Barcode Only) Feature Reference CSV file |
* HDF5 files now contain indices of the targeted genes within the features HDF5 group.