DropletQC: improved identification of empty droplets and damaged cells in single-cell RNA-seq data
Advances in droplet-based single cell RNA-sequencing (scRNA-seq) have dramatically increased throughput, allowing tens of thousands of cells to be routinely sequenced in a single experiment. In addition to cells, droplets capture cell-free "ambient" RNA predominately caused by lysis of cells during sample preparation. Samples with high ambient RNA concentration can create challenges in accurately distinguishing cell-containing droplets and droplets containing ambient RNA. Current methods to separate these groups often retain a significant number of droplets that do not contain cells - so called empty droplets. Additional to the challenge of identifying empty drops, there are currently no methods available to detect droplets containing damaged cells, which comprise of partially lysed cells - the original source of the ambient RNA. Here we describe DropletQC, a new method that is able to detect empty droplets, damaged, and intact cells, and accurately distinguish from one another. This approach is based on a novel quality control metric, the nuclear fraction, which quantifies for each droplet the fraction of RNA originating from unspliced, nuclear pre-mRNA. We demonstrate how DropletQC provides a powerful extension to existing computational methods for identifying empty droplets such as EmptyDrops. We have implemented DropletQC as an R package, which can be easily integrated into existing single cell analysis workflows.