Summary: Advances in next generation sequencing have generated massive amounts of short reads. However, assembling genome sequences from short reads still remains a challenging task. Due to errors in reads and large repeats in the genome, many of current assembly tools usually produce just collections of contigs whose relative positions and orientations along the genome being sequenced are still unknown. To address this issue, a scaffolding process to order and orient the contigs of a draft genome is needed for completing the genome sequence. In this work, we propose a new scaffolding tool called CSAR that can efficiently and more accurately order and orient the contigs of a given draft genome based on a reference genome of a related organism. In particular, the reference genome required by CSAR is not necessary to be complete in sequence. Our experimental results on real datasets have shown that CSAR outperforms other similar tools such as Projector2, OSLay and Mauve Aligner in terms of average sensitivity, precision, F-score, genome coverage, NGA50 and running time. Availability and implementation: The program of CSAR can be downloaded from https://github.com/ablab-nthu/CSAR. Contact: [email protected] or [email protected]. Supplementary information: Supplementary data are available at Bioinformatics online.

BACKGROUND: A draft genome assembled by current next-generation sequencing techniques from short reads is just a collection of contigs, whose relative positions and orientations along the genome being sequenced are unknown. To further obtain its complete sequence, a contig scaffolding process is usually applied to order and orient the contigs in the draft genome. Although several single reference-based scaffolding tools have been proposed, they may produce erroneous scaffolds if there are rearrangements between the target and reference genomes or their phylogenetic relationship is distant. This may suggest that a single reference genome may not be sufficient to produce correct scaffolds of a draft genome. RESULTS: In this study, we design a simple heuristic method to further revise our single reference-based scaffolding tool CAR into a new one called Multi-CAR such that it can utilize multiple complete genomes of related organisms as references to more accurately order and orient the contigs of a draft genome. In practical usage, our Multi-CAR does not require prior knowledge concerning phylogenetic relationships among the draft and reference genomes and libraries of paired-end reads. To validate Multi-CAR, we have tested it on a real dataset composed of several prokaryotic genomes and also compared its accuracy performance with other multiple reference-based scaffolding tools Ragout and MeDuSa. Our experimental results have finally shown that Multi-CAR indeed outperforms Ragout and MeDuSa in terms of sensitivity, precision, genome coverage, scaffold number and scaffold N50 size. CONCLUSIONS: Multi-CAR serves as an efficient tool that can more accurately order and orient the contigs of a draft genome based on multiple reference genomes. The web server of Multi-CAR is freely available at http://genome.cs.nthu.edu.tw/Multi-CAR/ .