Abstract
The availability of complete microbial genome sequences enormously facilitates experimental molecular investigations of the respective organisms by providing complete lists of genes, their genetic contexts, and their predicted functions. This can be used in a number of ways to focus studies on bacterial pathogenesis and also vaccine development (1,2). The complete genome sequences from two unrelated strains of Neisseria meningitidis, a derivative of isolate MC58 which originally expressed serogroup B capsule and strain Z2491, which is serogroup A, are now available (3,4). The genome sequences of both these strains were determined using the whole genome shotgun approach (5). In this approach, randomly sheared chromosomal DNA is cloned to make a small insert library (1.5–2.0 kb for MC58, 0.5–0.8 kb and 1.0–1.5 kb for Z2491), then each insert is sequenced from both ends using plasmidspecific primers. For the MC58 genome sequence, a large insert lambda library (8–24 kb) was also used. In the initial sequencing phase, 6-8 times coverage of the estimated size of the genome is generally achieved. The DNA sequences are linked together (assembled) into large contigs (a derivative of the word contiguous). Polymerase chain reaction (PCR) and sequencing of large insert libraries are then used to join the contigs, close gaps, and resolve ambiguities (see ref. 6 for a review).
This is a preview of subscription content, log in via an institution.
Buying options
Tax calculation will be finalised at checkout
Purchases are for personal use only
Learn about institutional subscriptionsReferences
Saunders N. J. and Moxon E. R. (1998) Implications of sequencing bacterial genomes for pathogenesis and vaccine development. Curr. Opin. Biotechnol. 9, 618–623.
Field D., Hood D., and Moxon E. R. (1999) Contribution of genomics to bacterial pathogenesis. Curr. Opin. Genet. Dev. 9, 700–703.
Tettelin H., Saunders N. J., Heidelberg J., Jeffries A. C., Nelson K. E., Eisen J. A., et al. (dy2000) Complete genome sequence of Neisseria meningitidis serotype B strain MC58. Science 287, 1809–1815.
Parkhill J., Achtman M., James K. D., Bentley S. D., Churcher C., Klee S. R., et al. (dy2000) Complete DNA sequence of a serogroup A strain of Neisseria meningitidis Z2491. Nature 404, 502–506.
Fleischmann R. D., Adams M. D., White O., Clayton R. A., Kirkness E. F., Kerlavage A. R., et al. (dy1995) Whole-genome random sequencing and assembly of Haemophilus influenzae Rd. Science 269, 496–512.
Frangeul L., Nelson K. E., Buchrieser C., Danchin A., Glaser P., and Kunst F. (1999) Cloning and assembly strategies in microbial genome projects. Microbiology 145, 2625–2634.
Salzberg S., Delcher A., Kasif S., and White O. (1998) Microbial gene identification using interpolated Markov models. Nucleic Acids Res. 2, 544–548.
Saunders N. J., Peden J. F., Hood D. W., and Moxon E. R. (1998) Simple sequence repeats in the Helicobacter pylori genome. Mol. Microbiol. 27, 1091–1098.
Moxon E. R., Rainey P. B., Nowak M. A., and Lenski R. E. (1994) Adaptive evolution of highly mutable loci in pathogenic bacteria. Curr. Biol. 4, 24–33.
Saunders N. J., Jeffries A. C., Peden J. F., Hood D. W., Tettelin H., Rappuoli R. and Moxon E. R. (2000) Repeat-associated phase variable genes in the complete genome sequence of Neisseria meningitidis strain MC58. Mol. Micobiol. 37, 207–215.
Tatusov R. L., Koonin E. V., and Lipman D. J. (1997) A genomic perspective on protein families. Science 278, 631–63
Tatusov R. L., Galperin M. Y., Natale D. A., and Koonin E. V. (2000) The COG database: a tool for genome-scale analyses of protein functions and evolution. Nucleic Acids Res. 28, 33–36.
Altschul S. F., Gish W., Miller W., Myers E. W., and Lipman D. J. (1990) Basic local alignment search tool. J. Mol. Biol. 215, 403–410.
Altschul S. F., Madden T. L., Schaffer A. A., Zhang J., Zhang Z., Miller W., and Lipman D. J. (1997) Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 25, 3389–3402.
Pearson W. R. (2000) Flexible sequence similarity searching with the FASTA3 program package. Methods Mol. Biol. 132, 185–219.
Henikoff S. and Henikoff J. G. (1992) Amino acid substitution matrices from protein blocks. Proc. Natl. Acad. Sci. USA 89, 10,915–10,919.
Galperin M. Y. and Koonin E. V. (1998) Sources of systematic error in functional annotation of genomes: domain rearrangement non-orthologous gene displacement and operon disruption. In Silico Biol. 1, 55–67.
Enright A. J., Iliopoulos I., Kyprides N. C., and Ouzounis C. A. (1999) Protein interaction maps for complete genomes based on gene fusion events. Nature 402, 86–90.
Kyrpides N. C. and Ouzounis C. A. (1999) Whole-genome sequence annotation: ‘Going wrong with confidence.’ Mol. Microbiol. 32, 881–891.
Bateman A., Birney E., Durbin R., Eddy S. R., Howe K. L., and Sonnhammer E. L. L. (2000) The Pfam protein families database. Nucleic Acids Res. 28, 263–266.
Bucher P. and Bairoch A. (1994) A generalized profile syntax for biomolecular sequences motifs and its function in automatic sequence interpretation, in ISMB-94 Proceedings 2nd International Conference on Intelligent Systems for Molecular Biology (Altman R., Brutlag D., Karp P., Lathrop R., and Searls D., eds.), AAAI Press, Menlo Park, OH, pp. 53–61.
Corpet F., Gouzyl J., and Kahn D. (1999) Recent improvements of the ProDom database of protein domain families. Nucleic Acids Res. 27, 263–267.
Doolittle R. F. (1995) The multiplicity of domains in proteins. Ann. Rev. Biochem. 64, 287–314.
Kanehisa M. and Goto S. (2000) KEGG: Kyoto encyclopedia of genes and genomes. Nucleic Acids Res. 28, 27–30.
Galperin M. Y. and Koonin E. V. (1999) Functional genomics and genome evolution. Genetica 106, 159–170.
Galperin M. Y., Tatusov R. L., and Koonin E. V. (1999) Comparing microbial genomes: how the gene set determines the lifestyle, in Organization of the Prokary otic Genome (Charlebois R. L., ed.), ASM, Washington, DC, pp. 91–108.
Riley M. (1998) Systems for categorizing functions of gene products. Curr. Opin. Struct. Biol. 8, 388–392.
Koonin E. V., Mushegian A. R., and Boork P. (1996) Non-orthologous gene displacement. Trends Genet. 12, 334–336.
Henikoff S., Greene E. A., Pietrokovski S., Bork P., Attwood T. K., and Hood L. (1997) Gene families: the taxonomy of protein paralogs and chimeras. Science 278, 609–614.
Tomb J. F., White O., Kerlavage A. R., Clayton R. A., Sutton G. G., Fleischmann R. D., et al. (1997) The complete genome sequence of the gastric pathogen Helicobacter pylori. Nature 388, 539–547.
Overbeek R., Fonstein M., D’Souza M., Pusch G. D., and Malsev N. (1999) The use of gene clusters to infer functional coupling. Proc. Natl. Acad. Sci. USA 96, 2896–2901.
Ouellette B. F. F. (1998) The GenBank sequence database, in Bioinformatics:A Practical Guide to the Analysis of Genes and Proteins Baxevanis A. D. and Ouellette B. F. F., eds.), Wiley, New York, pp. 16–45.
Delcher A. L., Kasif S., Fleischmann R. D., Peterson J., White O., and Salzberg S. L. (1999) Alignment of whole genomes. Nucleic Acids Res. 27, 2369–2376.
Karlin S., Campbell A. M., and Mrazek J. (1998) Comparative DNA analysis across diverse genomes. Ann. Rev. Genet. 32, 185–225.
Pizza M., Scarlato V., Masignani V., Giuliani M. M., Arico B., Baldi L., et al. (dy2000) Novel proteins for vaccine development from the meningococcus B genome. Science 287, 1816–1820.
Saunders N. J., Peden J. F., and Moxon E. R. (1999) The absence of an uptake sequence in Helicobacter pylori. Microbiology 145, 3523–3528.
Hosking S. L., Deadman M. E., Moxon E. R., Peden J. F., Saunders N. J., and High N. J. (1998) An in silico evaluation of Tn916 as a tool for generalisedmutagenesis in Haemophilus influenzae Rd. Microbiology 144, 2525–2530.
Wilson M., DeRisi J., Kristensen H-H., Imboden P., Rane S., Brown P. O., and Schoolnik G. K. (1999) Exploring drug-induced alterations in gene expression by microarray hybridization. Proc. Natl. Acad. Sci. USA 96, 12,833–12,838.
Behr M. A., Wilson M. A., Gill W. P., Salamon H., Schoolnik G. K., Rane S., and Small P. M. (1999) Comparative genomics of BCG vaccines by whole-genome DNA microarray. Science 284, 1520–1523.
Wasinger V. C., Pollack J. D., and Humphery-Smith I. (2000) The proteome of Mycoplasma genitalium chaps-soluble component. Eur. J. Biochem. 267, 1571–1582.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2001 Humana Press Inc., Totowa, NJ
About this protocol
Cite this protocol
Jeffries, A.C., Saunders, N.J., Hood, D.W. (2001). Genome Sequencing and Annotation. In: Walker, J.M., Pollard, A.J., Maiden, M.C.J. (eds) Meningococcal Disease. Methods in Molecular Medicine™, vol 67. Humana Press. https://doi.org/10.1385/1-59259-149-3:215
Download citation
DOI: https://doi.org/10.1385/1-59259-149-3:215
Publisher Name: Humana Press
Print ISBN: 978-0-89603-849-3
Online ISBN: 978-1-59259-149-7
eBook Packages: Springer Protocols