Abstract
An important but rarely discussed phenomenon in single cell data generated by the 10X-Chromium protocol is that the fraction of non-exonic reads is very high. This number usually exceeds 30% of the total reads. Without aligning them to a complete genome reference, non-exonic reads can be erroneously aligned to the transcriptome reference with higher error rates. To tackle this problem, Cell Ranger chooses to firstly align reads against the whole genome, and at a later step, uses a genome annotation to select reads that align to the transcriptome. Despite its high running time and large memory consumption, Cell Ranger remains the most widely used tool to quantify 10XGenomics single cell RNA-Seq data for its accuracy.
In this work, we introduce Hera-T, a fast and accurate tool for estimating gene abundances in single cell data generated by the 10X-Chromium protocol. By devising a new strategy for aligning reads to both transcriptome and genome references, Hera-T reduces both running time and memory consumption from 10 to 100 folds while giving similar results compared to Cell Ranger’s. Hera-T also addresses some difficult splicing alignment scenarios that Cell Ranger fails to address, and therefore, obtains better accuracy compared to Cell Ranger. Excluding the reads in those scenarios, Hera-T and Cell Ranger results have correlation scores > 0.99.
For a single-cell data set with 49 million of reads, Cell Ranger took 3 hours (179 minutes) while Hera-T took 1.75 minutes; for another single-cell data set with 784 millions of reads, Cell Ranger took about 25 hours while Hera-T took 32 minutes. For those data sets, Cell Ranger completely used all 32 GB of memory while Hera-T consumed at most 8 GB. Hera-T package is available for download at: https://bioturing.com/product/hera-t
