Paired cloning vectors for complementation of mutations in the cyanobacterium<Emphasis Type="Italic"> Anabaena</Emphasis> sp. strain PCC 7120 |
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Authors: | C Peter Wolk Qing Fan Ruanbao Zhou Guocun Huang Sigal Lechno-Yossef Tanya Kuritz Elizabeth Wojciuch |
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Institution: | (1) MSU-DOE Plant Research Laboratory, Michigan State University, East Lansing, MI 48824-1312, USA;(2) Department of Plant Biology, Michigan State University, East Lansing, MI 48824-1312, USA;(3) Present address: Feinberg School of Medicine, Northwestern University, 303 East Chicago Avenue, Chicago, IL 60611-3008, USA;(4) Present address: College of Life Sciences, Anhui Normal University, Wuhu, Anhui, 241000, People’s Republic of China;(5) Present address: Department of Physiology, University of Texas Southwestern Medical Center at Dallas, 5323 Harry Hines Boulevard, Dallas, TX 75390, USA;(6) Present address: Department of Chemistry, Michigan State University, East Lansing, MI 48824-1312, USA;(7) Present address: Chemical Sciences Division, Oak Ridge National Laboratory, MS-6194, Oak Ridge, TN 37830, USA |
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Abstract: | 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. |
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Keywords: | Anabaena sp strain PCC 7120 Cloning vectors Complementation of mutations |
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