Validating Paired-End Read Alignments in Sequence Graphs

Graph based non-linear reference structures such as variation graphs and colored de Bruijn graphs enable incorporation of full genomic diversity within a population. However, transitioning from a simple string-based reference to graphs requires addressing many computational challenges, one of which concerns accurately mapping sequencing read sets to graphs. Paired-end Illumina sequencing is a commonly used sequencing platform in genomics, where the paired-end distance constraints allow disambiguation of repeats. Many recent works have explored provably good index-based and alignment-based strategies for mapping individual reads to graphs. However, validating distance constraints efficiently over graphs is not trivial, and existing sequence to graph mappers rely on heuristics. We introduce a mathematical formulation of the problem, and provide a new algorithm to solve it exactly. We take advantage of the high sparsity of reference graphs, and use sparse matrix-matrix multiplications (SpGEMM) to build an index which can be queried efficiently by a mapping algorithm for validating the distance constraints. Effectiveness of the algorithm is demonstrated using real reference graphs, including a human MHC variation graph, and a pan-genome de-Bruijn graph built using genomes of 20 B. anthracis strains. While the one-time indexing time can vary from a few minutes to a few hours using our algorithm, answering a million distance queries takes less than a second

Near-optimal RNA-Seq quantification

We present a novel approach to RNA-Seq quantification that is near optimal in speed and accuracy. Software implementing the approach, called kallisto, can be used to analyze 30 million unaligned paired-end RNA-Seq reads in less than 5 minutes on a standard laptop computer while providing results as accurate as those of the best existing tools. This removes a major computational bottleneck in RNA-Seq analysis.Comment: - Added some results (paralog analysis, allele specific expression analysis, alignment comparison, accuracy analysis with TPMs) - Switched bootstrap analysis to human sample from SEQC-MAQCIII - Provided link to a snakefile that allows for reproducibility of all results and figures in the pape

Exploring single-sample SNP and INDEL calling with whole-genome de novo assembly

Motivation: Eugene Myers in his string graph paper (Myers, 2005) suggested that in a string graph or equivalently a unitig graph, any path spells a valid assembly. As a string/unitig graph also encodes every valid assembly of reads, such a graph, provided that it can be constructed correctly, is in fact a lossless representation of reads. In principle, every analysis based on whole-genome shotgun sequencing (WGS) data, such as SNP and insertion/deletion (INDEL) calling, can also be achieved with unitigs. Results: To explore the feasibility of using de novo assembly in the context of resequencing, we developed a de novo assembler, fermi, that assembles Illumina short reads into unitigs while preserving most of information of the input reads. SNPs and INDELs can be called by mapping the unitigs against a reference genome. By applying the method on 35-fold human resequencing data, we showed that in comparison to the standard pipeline, our approach yields similar accuracy for SNP calling and better results for INDEL calling. It has higher sensitivity than other de novo assembly based methods for variant calling. Our work suggests that variant calling with de novo assembly be a beneficial complement to the standard variant calling pipeline for whole-genome resequencing. In the methodological aspects, we proposed FMD-index for forward-backward extension of DNA sequences, a fast algorithm for finding all super-maximal exact matches and one-pass construction of unitigs from an FMD-index. Availability: http://github.com/lh3/fermi Contact: [email protected]: Rev2: submitted version with minor improvements; 7 page