The Single-cell QA/QC task in Partek Flow enables you to visualize several measure of cell quality and filter to include only high-quality cells. To invoke Single-cell QA/QC: 

• Click a Single cell counts data node
• Click the QA/QC section of the task menu
• Click Single cell QA/QC

If your Single cell counts data node has been annotated with a gene/transcript annotation, the task will run without a task configuration dialog. However, if you imported a single cell counts matrix without specifying an gene/transcript annotation file, you will be prompted to choose the genome assembly and annotation file by the Single cell QA/QC configuration dialog (Figure 1). 

 

Figure 1. If an annotation was not specified on import, you can choose the assembly and annotation after invoking Single-cell QA/QC

The Single cell QA/QC task report includes interactive violin plots showing the value of every cell in the project on two or three quality measures (Figure 2).

 

Figure 2. Interactive violin plots showing values of each cell for several quality measures

There are typically three plots: counts per cell, detected genes per cell, and the percentage of mitochondrial counts per cell. If your cells do not express any mitochondrial genes or the IDs in the input file do not match our expected IDs for mitochondrial genes, the plot for the percentage of mitochondrial counts per cell will be absent. 

Counts is calculated as the sum of the counts for all features in each cell from the input data node. Detected genes is calculated as the number of features in each cell with greater than zero counts. Percentage of mitochondrial counts is calculated as the sum of counts for known mitochondrial genes divided by the sum of counts for all features and multiplied by 100. 

Each point on the plots is a cell. All cells from all samples are shown on the plots. The pink violins illustrate the distribution of cell values for the y-axis metric. 

There are two methods for filtering cells. First, cells can be filtered by clicking and dragging to select a range on one of the plots (Figure 3)

 

The selected region remains bright while the excluded region is shaded. Selected cells are included and cells that are not selected are excluded. Cell that are excluded turn black. The plots are linked so that cells excluded on one plot are shown as excluded on all plots (Figure 4). The filters are additive; combining multiple filters will include the intersection of the three filters. 

 

Figure 4. Selected cells are shown in blue, excluded cells are shown in black

Alternatively, the filters can be set using the text boxes below each plot (Figure 5). The minimum and maximum of the filter can be set using Counts or Percentiles for the Counts filter and Detected genes filters. For the Mitochondrial counts filter, you can set the minimum and maximum mitochondrial reads percentage.  The number and percentage of cells included in the filter is listed at the bottom of the page and updates as filters are added. 

 

Figure 5. Setting the max and min of each filter using the text boxes

It can be helpful to view the range of values for Counts and Detected genes on a log scale. To switch the y-axis of these plots to a log scale, click the checkbox at the top of the page (Figure 6)

 

Figure 6. Log scale the y-axis of the counts and detected genes plots

For data sets with very many cells, it may be helpful to decrease the dot opacity to better visualize the plot density. Dot opacity can be adjusted using the slider at the top of the page (Figure 7).

 

To apply the filter and generate a filtered version of the input data node, click Apply filter.  

A new data node, Filtered single cell counts, will be generated (Figure 8).

 

Figure 8. Filter cells task runs from the Single cell QA/QC report

The Filter cells task report includes the filter criteria, lists the feature distribution statistics for each sample, and gives a breakdown of how many and what percentage of cells were excluded from each sample by each filter (Figure 9). 

 

Figure 9. The filter cells task report gives filter settings and a breakdown of filtering by sample

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