10x Genomics
Chromium Single Cell Gene Expression

Cell Ranger3.0, printed on 11/21/2024

Run Summary

The cellranger pipeline outputs a summary HTML file named web_summary.html that contains summary metrics and automated secondary analysis results. Rarely, an issue may also be detected during the pipeline run leading to an alert also appearing on this page, and more information about these is provided in the troubleshooting documentation.

Summary View

The run summary can be viewed by clicking "Summary" in the top left corner. The summary metrics describe sequencing quality and various characteristics of the detected cells.

The number of cells detected, the mean reads per cell, and the median genes detected per cell are prominently displayed near the top of the page.

Click the '?' in the upper right corner of each dashboard for more information on each metric.

The Barcode Rank Plot under the "Cells" dashboard shows the distribution of barcode counts and which barcodes were inferred to be associated with cells. The y-axis is the number of UMI counts mapped to each barcode and the x-axis is the number of barcodes below that value. A steep drop-off is indicative of good separation between the cell-associated barcodes and the barcodes associated with empty partitions. Barcodes can be determined to be cell-associated based on their UMI count or by their RNA profiles, therefore some regions of the graph can contain both cell-associated and background-associated barcodes. The color of the graph represents the local density of barcodes that are cell-associated. See Cell Calling Algorithms for more details.

Analysis View

The automated secondary analysis results can be viewed by clicking "Analysis" in the top left corner. The secondary analysis provides the following:

• A dimensional reduction analysis which projects the cells into a 2-D space (t-SNE)
• An automated clustering analysis which groups together cells that have similar expression profiles
• A list of genes that are differentially expressed between the selected clusters
• A plot showing the effect of decreased sequencing depth on observed library complexity
• A plot showing the effect of decreased sequencing depth on median genes per cell detected

The top left plot shows the 2-D t-SNE projection of the cells colored by the total UMI counts per cell. This is suggestive of the RNA content of the cells and often correlates with cell size - redder points are cells with more RNA in them.

The top right plot overlays the clustering results onto the 2-D t-SNE projection of cells. The type of clustering analysis is selectable from the dropdown in the upper right - change this to vary the type of clustering and/or number of clusters that are assigned to the data.

The table in the middle shows which genes are differentially expressed in each cluster relative to all other clusters. To find the genes associated with a particular cluster, you can click the cluster number to sort the table by specificity for that cluster.

The bottom left plot shows the effect of decreased sequencing depth on Sequencing Saturation, which is a measure of the fraction of library complexity that was observed. The far right point of the line is the full sequencing depth obtained in this run.

Similarly, the bottom right plot shows the effect of decreased sequencing depth on Median Genes per Cell, which is a way of measuring data yield as a function of depth. The far right point is the full sequencing depth obtained in this run.

Feature Barcoding View

Cell Ranger 3.0 introduced Feature Barcoding analysis. If you have Cell Ranger with Feature Barcoding libraries, new metrics that are specific to the given Feature Barcoding libraries will appear in summary tab.

Below is an example web summary when the Antibody Capture library type is used:

Here is an example web summary when the CRISPR Guide Capture library type is used:

The analysis view for Feature Barcoding experiments contains the same secondary analysis as purely gene expression experiments, and uses only the gene expression part of the feature-barcode matrix for dimensionality reduction, clustering, and differential gene expression analysis.

Multiple Species

If you analyzed a multi-species experiment, the output will look different. For example, the human-mouse mixing experiment is run to verify system functionality. It consists of mixing approximately 500 human (HEK293T) cells and 500 mouse (3T3) cells in a 1:1 ratio.

Click the '?' in the upper right corner of each dashboard for more information on each metric.

In the Analysis View, running the human-mouse mixing experiment results in the following plot:

Each point represents a barcode; the gray points are inferred to be associated with GEM partitions that contained more than one cell because they contained a large number of molecules that came from human cells and mouse cells. The GEMs containing human-human and mouse-mouse cell combinations are not visible here, but their presence is inferred in the "Fraction GEMs with >1 Cell" metric in the Summary View.