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A hybrid approach for the automated finishing of bacterial genomes.

ABSTRACT: Advances in DNA sequencing technology have improved our ability to characterize most genomic diversity. However, accurate resolution of large structural events is challenging because of the short read lengths of second-generation technologies. Third-generation sequencing technologies, which can yield longer multikilobase reads, have the potential to address limitations associated with genome assembly. Here we combine sequencing data from second- and third-generation DNA sequencing technologies to assemble the two-chromosome genome of a recent Haitian cholera outbreak strain into two nearly finished contigs at >99.9% accuracy. Complex regions with clinically relevant structure were completely resolved. In separate control assemblies on experimental and simulated data for the canonical N16961 cholera reference strain, we obtained 14 scaffolds of greater than 1 kb for the experimental data and 8 scaffolds of greater than 1 kb for the simulated data, which allowed us to correct several errors in contigs assembled from the short-read data alone. This work provides a blueprint for the next generation of rapid microbial identification and full-genome assembly.

SUBMITTER: Bashir A

PROVIDER: S-EPMC3731737 | biostudies-literature | 2012 Jul

REPOSITORIES: biostudies-literature

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A hybrid approach for the automated finishing of bacterial genomes.

Bashir Ali A Klammer Aaron A Robins William P WP Chin Chen-Shan CS Webster Dale D Paxinos Ellen E Hsu David D Ashby Meredith M Wang Susana S Peluso Paul P Sebra Robert R Sorenson Jon J Bullard James J Yen Jackie J Valdovino Marie M Mollova Emilia E Luong Khai K Lin Steven S LaMay Brianna B Joshi Amruta A Rowe Lori L Frace Michael M Tarr Cheryl L CL Turnsek Maryann M Davis Brigid M BM Kasarskis Andrew A Mekalanos John J JJ Waldor Matthew K MK Schadt Eric E EE

Nature biotechnology 20120701 7

Advances in DNA sequencing technology have improved our ability to characterize most genomic diversity. However, accurate resolution of large structural events is challenging because of the short read lengths of second-generation technologies. Third-generation sequencing technologies, which can yield longer multikilobase reads, have the potential to address limitations associated with genome assembly. Here we combine sequencing data from second- and third-generation DNA sequencing technologies t ...[more]

PMID: 22750883

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Project description:Currently, the genome sequencing community is producing shotgun sequence data at a very high rate, but finishing (collecting additional directed sequence data to close gaps and improve the quality of the data) is not matching that rate. One reason for the difference is that shotgun sequencing is highly automated but finishing is not: Most finishing decisions, such as which directed reads to obtain and which specialized sequencing techniques to use, are made by people. If finishing rates are to increase to match shotgun sequencing rates, most finishing decisions also must be automated. The Autofinish computer program (which is part of the computer software package) does this by automatically choosing finishing reads. Autofinish is able to suggest most finishing reads required for completion of each sequencing project, greatly reducing the amount of human attention needed. sometimes completely finishes the project, with no human decisions required. It cannot solve the most complex problems, so we recommend that Autofinish be allowed to suggest reads for the first three rounds of finishing, and if the project still is not finished completely, a human finisher complete the work. We compared this Autofinish-Hybrid method of finishing against a human finisher in five different projects with a variety of shotgun depths by finishing each project twice--once with each method. This comparison shows that the Autofinish-Hybrid method saves many hours over a human finisher alone, while using roughly the same number and type of reads and closing gaps at roughly the same rate. Autofinish currently is in production use at several large sequencing centers. It is designed to be adaptable to the finishing strategy of the lab--it can finish using some or all of the following: resequencing reads, reverses, custom primer walks on either subclone templates or whole clone templates, PCR, or minilibraries. Autofinish has been used for finishing cDNA, genomic clones, and whole bacterial genomes (see http://www.phrap.org).

Dataset Information

A hybrid approach for the automated finishing of bacterial genomes.

Publications

A hybrid approach for the automated finishing of bacterial genomes.

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