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Normalization of single-cell RNA sequencing data is necessary to eliminate cell-specific biases prior to downstream analyses. However, this is not straightforward for noisy single-cell data where many counts are zero. We present a novel approach where expression values are summed across pools of cells, and the summed values are used for normalization. Pool-based size factors are then deconvolved to yield cell-based factors. Our deconvolution approach outperforms existing methods for accurate normalization of cell-specific biases in simulated data. Similar behavior is observed in real data, where deconvolution improves the relevance of results of downstream analyses.   Electronic supplementary material  The online version of this article (doi:10.1186/s13059-016-0947-7) contains supplementary material, which is available to authorized users.

Pooling across cells to normalize single-cell RNA sequencing data with many zero counts