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The biological reduction of nitrogen gas to ammonia is limited to a select group of nitrogen-fixing prokaryotes. While nitrogenase is the catalyst of nitrogen fixation in these biological systems, a consortium of additional gene products is required for the synthesis, activation, and catalytic competency of this oxygen-sensitive metalloenzyme. Thus, the biochemical complexity of this process often requires functional studies and isolation of gene products from the native nitrogen-fixing organisms. The strict aerobe Azotobacter vinelandii is the best-studied model bacterium among diazotrophs. This chapter provides a description of procedures for targeted genomic manipulation and isolation of A. vinelandii strains. These methods have enabled identification and characterization of gene products with roles in nitrogen fixation and other related aspects of metabolism. The ability to modify and control expression levels of targeted sequences provides a biotechnological tool to uncover molecular details associated with nitrogen fixation, as well as to exploit this model system as a host for expression of oxygen-sensitive proteins.
This article was published in the following journal.
Name: Methods in molecular biology (Clifton, N.J.)
Azotobacter vinelandii produces differentiated cells, called cysts, surrounded by two alginate layers, which are necessary for their desiccation resistance. This alginate contains variable proportions...
A major hurdle in the studies of nitrogenase, one of the most complicated metalloenzymes known to date, is to obtain large amounts of intact, active proteins. Nitrogenase and related proteins are ofte...
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A species of gram-negative, aerobic bacteria first isolated from soil in Vineland, New Jersey. Ammonium and nitrate are used as nitrogen sources by this bacterium. It is distinguished from other members of its genus by the ability to use rhamnose as a carbon source. (From Bergey's Manual of Determinative Bacteriology, 9th ed)
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Contiguous large-scale (1000-400,000 basepairs) differences in the genomic DNA between individuals, due to SEQUENCE DELETION; SEQUENCE INSERTION; or SEQUENCE INVERSION.
The systematic study of annotated genomic information to global protein expression in order to determine the relationship between genomic sequences and both expressed proteins and predicted protein sequences.
A method for analyzing and mapping differences in the copy number of specific genes or other large sequences between two sets of chromosomal DNA. It is used to look for large sequence changes such as deletions, duplications, or amplifications within the genomic DNA of an individual (with a tumor for example) or family members or population or between species.
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