title: Strategy to sequence the genome of Corynebacterium glutamicum ATCC 13032: use of a cosmid and a bacterial artificial chromosome library.
abstract: The initial strategy of the Corynebacterium glutamicum genome project was to sequence overlapping inserts of an ordered cosmid library. High-density colony grids of approximately 28 genome equivalents were used for the identification of overlapping clones by Southern hybridization. Altogether 18 contiguous genomic segments comprising 95 overlapping cosmids were assembled. Systematic shotgun sequencing of the assembled cosmid set revealed that only 2.84 Mb (86.6%) of the C. glutamicum genome were represented by the cosmid library. To obtain a complete genome coverage, a bacterial artificial chromosome (BAC) library of the C. glutamicum chromosome was constructed in pBeloBAC11 and used for genome mapping. The BAC library consists of 3168 BACs and represents a theoretical 63-fold coverage of the C. glutamicum genome (3.28 Mb). Southern screening of 2304 BAC clones with PCR-amplified chromosomal markers and subsequent insert terminal sequencing allowed the identification of 119 BACs covering the entire chromosome of C. glutamicum. The minimal set representing a 100% genome coverage contains 44 unique BAC clones with an average overlap of 22 kb. A total of 21 BACs represented linking clones between previously sequenced cosmid contigs and provided a valuable tool for completing the genome sequence of C. glutamicum.
authors.affiliation: Zentrum für Genomforschung, Universität Bielefeld, Universitätsstrasse 25, D-33615 Bielefeld, Germany. andreas.tauch@genetik.uni-bielefeld.de
authors.forename: Andreas
authors.initials: A
authors.lastname: Tauch
authors.forename: Iris
authors.initials: I
authors.lastname: Homann
authors.forename: Sascha
authors.initials: S
authors.lastname: Mormann
authors.forename: Silvia
authors.initials: S
authors.lastname: Rüberg
authors.forename: Alain
authors.initials: A
authors.lastname: Billault
authors.forename: Brigitte
authors.initials: B
authors.lastname: Bathe
authors.forename: Sven
authors.initials: S
authors.lastname: Brand
authors.forename: Olaf
authors.initials: O
authors.lastname: Brockmann-Gretza
authors.forename: Christian
authors.initials: C
authors.lastname: Rückert
authors.forename: Natalie
authors.initials: N
authors.lastname: Schischka
authors.forename: Carsten
authors.initials: C
authors.lastname: Wrenger
authors.forename: Jörg
authors.initials: J
authors.lastname: Hoheisel
authors.forename: Bettina
authors.initials: B
authors.lastname: Möckel
authors.forename: Klaus
authors.initials: K
authors.lastname: Huthmacher
authors.forename: Walter
authors.initials: W
authors.lastname: Pfefferle
authors.forename: Alfred
authors.initials: A
authors.lastname: Pühler
authors.forename: Jörn
authors.initials: J
authors.lastname: Kalinowski
mesh_terms: D022202:Chromosomes, Artificial, Bacterial / Q000235:genetics*; D002876:Chromosomes, Bacterial / Q000235:genetics*; D003001:Cloning, Molecular; D020451:Contig Mapping / Q000379:methods*; D003352:Corynebacterium / Q000235:genetics*; D003360:Cosmids / Q000235:genetics*; D005798:Genes, Bacterial / Q000235:genetics; D016680:Genome, Bacterial; D015698:Genomic Library; D023281:Genomics / Q000379:methods; D017422:Sequence Analysis, DNA / Q000379:methods