OpenSSL SSL_connect: Connection reset by peer in connection to v2.sherpa.ac.uk:443 PuSH - Publication Server of Helmholtz Zentrum München: <em>De novo</em> 454 sequencing of barcoded BAC pools for comprehensive gene survey and genome analysis in the complex genome of barley.

PuSH - Publication Server of Helmholtz Zentrum München

Steuernagel, B.* ; Taudien, S.* ; Gundlach, H. ; Seidel, M. ; Ariyadasa, R.* ; Schulte, D.* ; Petzold, A.* ; Felder, M.* ; Graner, A.* ; Scholz, U.* ; Mayer, K.F.X. ; Platzer, M.* ; Stein, N.*

De novo 454 sequencing of barcoded BAC pools for comprehensive gene survey and genome analysis in the complex genome of barley.

BMC Genomics 10:547 (2009)
Publ. Version/Full Text Volltext DOI PMC
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BACKGROUND: De novo sequencing the entire genome of a large complex plant genome like the one of barley (Hordeum vulgare L.) is a major challenge both in terms of experimental feasibility and costs. The emergence and breathtaking progress of next generation sequencing technologies has put this goal into focus and a clone based strategy combined with the 454/Roche technology is conceivable. RESULTS: To test the feasibility, we sequenced 91 barcoded, pooled, gene containing barley BACs using the GS FLX platform and assembled the sequences under iterative change of parameters. The BAC assemblies were characterized by N50 of approximately 50 kb (N80 approximately 31 kb, N90 approximately 21 kb) and a Q40 of 94%. For approximately 80% of the clones, the best assemblies consisted of less than 10 contigs at 24-fold mean sequence coverage. Moreover we show that gene containing regions seem to assemble completely and uninterrupted thus making the approach suitable for detecting complete and positionally anchored genes.By comparing the assemblies of four clones to their complete reference sequences generated by the Sanger method, we evaluated the distribution, quality and representativeness of the 454 sequences as well as the consistency and reliability of the assemblies. CONCLUSION: The described multiplex 454 sequencing of barcoded BACs leads to sequence consensi highly representative for the clones. Assemblies are correct for the majority of contigs. Though the resolution of complex repetitive structures requires additional experimental efforts, our approach paves the way for a clone based strategy of sequencing the barley genome.
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Publication type Article: Journal article
Document type Scientific Article
Corresponding Author
Keywords ARABIDOPSIS-THALIANA; DNA; IDENTIFICATION; EVOLUTION; SOFTWARE; CLONES; READS; TOOL
ISSN (print) / ISBN 1471-2164
e-ISSN 1471-2164
Journal BMC Genomics
Quellenangaben Volume: 10, Issue: , Pages: , Article Number: 547 Supplement: ,
Publisher BioMed Central
Non-patent literature Publications
Reviewing status Peer reviewed
Institute(s) Institute of Bioinformatics and Systems Biology (IBIS)