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. 2008 Dec 15;24(24):2818-24.
doi: 10.1093/bioinformatics/btn548. Epub 2008 Oct 24.

Aggressive assembly of pyrosequencing reads with mates

Jason R Miller  1 , Arthur L Delcher, Sergey Koren, Eli Venter, Brian P Walenz, Anushka Brownley, Justin Johnson, Kelvin Li, Clark Mobarry, Granger Sutton
Affiliations

Aggressive assembly of pyrosequencing reads with mates

Jason R Miller et al. Bioinformatics. .

Abstract

Motivation: DNA sequence reads from Sanger and pyrosequencing platforms differ in cost, accuracy, typical coverage, average read length and the variety of available paired-end protocols. Both read types can complement one another in a 'hybrid' approach to whole-genome shotgun sequencing projects, but assembly software must be modified to accommodate their different characteristics. This is true even of pyrosequencing mated and unmated read combinations. Without special modifications, assemblers tuned for homogeneous sequence data may perform poorly on hybrid data.

Results: Celera Assembler was modified for combinations of ABI 3730 and 454 FLX reads. The revised pipeline called CABOG (Celera Assembler with the Best Overlap Graph) is robust to homopolymer run length uncertainty, high read coverage and heterogeneous read lengths. In tests on four genomes, it generated the longest contigs among all assemblers tested. It exploited the mate constraints provided by paired-end reads from either platform to build larger contigs and scaffolds, which were validated by comparison to a finished reference sequence. A low rate of contig mis-assembly was detected in some CABOG assemblies, but this was reduced in the presence of sufficient mate pair data.

Availability: The software is freely available as open-source from http://wgs-assembler.sf.net under the GNU Public License.

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Figures

Fig. 1.
Fig. 1.
Two representations of a best overlap graph. In (a), the layout resembles a multiple sequence alignment. In (b) each read is represented by two nodes joined by an undirected edge. Arrows represent best overlaps, where best means covering the most sequence. There are mutual best overlaps between successive pairs of reads A through D. Due to erroneous bases at one end (wavy line), read E has a non-mutual best overlap to B. Paths span undirected and directed edges alternately. Path EBA converges on path ABCD. CABOG scores read E lower than the others since only three reads are on paths from it. Starting with any one of the high-scoring reads, CABOG would build initial unitig ABCD, then E. Using saved information about each path intersection, CABOG would discount the intersection at B because the path from E spanned only one read before B. It would break ABCD only if there were also a change in read arrival rate at B, which is not the case here. Although linear-time directed-path following finds the longest possible unitig in this constructed case, it is not guaranteed to do so when paths span multiple intersections.
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