Droplet-based microfluidic devices have become widely used to perform single-cell RNA sequencing (scRNA- seq) and discover novel cellular heterogeneity in complex biological systems. However, ambient RNA present in the cell suspension can be incorporated into these droplets and aberrantly counted along with a cell's native mRNA. This results in cross-contamination of transcripts between different cell populations and can potentially decrease the precision of downstream analyses. We developed a novel hierarchical Bayesian method called DecontX to estimate and remove contamination in individual cells from scRNA- seq data. DecontX accurately predicted the proportion of contaminated counts in a mixture of mouse and human cells. Decontamination of PBMC datasets removed aberrant expression of cell type specific marker genes from other cell types and improved overall separation of cell clusters. In general, DecontX can be incorporated into scRNA-seq workflows to assess quality of dissociation protocols and improve downstream analyses.

Decontamination of ambient RNA in single-cell RNA-seq with DecontX