Paired cloning vectors for complementation of mutations in the cyanobacterium Anabaena sp. strain PCC 7120

The clones generated in a sequencing project represent a resource for subsequent analysis of the organism whose genome has been sequenced. We describe an interrelated group of cloning vectors that either integrate into the genome or replicate, and that enhance the utility, for developmental and othe...

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Bibliographic Details
Published in:Archives of microbiology 2007-12, Vol.188 (6), p.551-563
Main Authors: Wolk, C. Peter, Fan, Qing, Zhou, Ruanbao, Huang, Guocun, Lechno-Yossef, Sigal, Kuritz, Tanya, Wojciuch, Elizabeth
Format: Article
Language:English
Subjects:
Anabaena
Anabaena - classification
Anabaena - genetics
Anabaena sp. strain PCC 7120
Bacteria
Bacteriology
BASIC BIOLOGICAL SCIENCES
Biological and medical sciences
CLONING
Cloning, Molecular
Complementation of mutations
Cyanophyta
DNA Replication - genetics
DNA Transposable Elements - genetics
Escherichia coli Proteins - genetics
Fundamental and applied biological sciences. Psychology
GENES
Genetic Complementation Test
genetic vectors
Genetic Vectors - genetics
Microbiology
Miscellaneous
Mutation
MUTATIONS
PLASMIDS
Plasmids - genetics
PROMOTERS
STRAINS
TRANSPOSONS
VECTORS
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Summary:The clones generated in a sequencing project represent a resource for subsequent analysis of the organism whose genome has been sequenced. We describe an interrelated group of cloning vectors that either integrate into the genome or replicate, and that enhance the utility, for developmental and other studies, of the clones used to determine the genomic sequence of the cyanobacterium, Anabaena sp. strain PCC 7120. One integrating vector is a mobilizable BAC vector that was used both to generate bridging clones and to complement transposon mutations. Upon addition of a cassette that permits mobilization and selection, pUC-based sequencing clones can also integrate into the genome and thereupon complement transposon mutations. The replicating vectors are based on cyanobacterial plasmid pDU1, whose sequence we report, and on broad-host-range plasmid RSF1010. The RSF1010- and pDU1-based vectors provide the opportunity to express different genes from either cell-type-specific or -generalist promoters, simultaneously from different plasmids in the same cyanobacterial cells. We show that pDU1 ORF4 and its upstream region play an essential role in the replication and copy number of pDU1, and that ORFs alr2887 and alr3546 (hetF A ) of Anabaena sp. are required specifically for fixation of dinitrogen under oxic conditions.
ISSN:0302-8933
1432-072X
DOI:10.1007/s00203-007-0276-z