How Long to Read Analysis of a Gene Cluster that is Responsible for CGMP Production by the Nitrogen Fixing Symbiont Azospirillum Brasilense

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Description

The plant associating bacterium Azospirillum brasilense makes mutually beneficial relationships with plants by performing nitrogen fixation and improving root development with secretion of various plant hormones including auxin. The interactions between plants and microbes are often mediated by signaling molecules. One of the signaling molecules secreted and synthesized by bacterial species Azospirillum brasilense is cyclic guanosine monophosphate (cGMP). Synthetic cGMP can induce root development and increase auxin signaling in Arabidopsis thaliana. The genes involved in cGMP synthesis are gcyA, gcyB, gcyC, gcyX, gcyX2, and cgrA. gcyA codes for a main catalytic unit of guanylyl cyclase; cgrA encodes CgrA, a cGMP binding positive regulator, and the function of other genes are not yet known. My hypothesis was that A. brasilense utilizes cGMP to improve root development of plants. To test this hypothesis, I examined whether the deletion of cGMP synthesis in A. brasilense causes disruptions in cGMP production and regulation. Toward this goal, I disrupted genes in gcy operon: gcyA, gcyB, gcyC, gcyX, gcyX2, and cgrA in A. brasilense. Each mutant was assayed with cGMP ELISA to check cGMP levels. The cGMP levels of all other mutants except gcyX were either close to zero or extremely low compared to the wildtype, whereas the cGMP level of the gcyX mutant was significantly higher than that of wildtype. To test if GcyX is a negative regulator of the gcy gene cluster, I performed qRT-PCR to determine the level of gcyA expression. gcyX mutant indeed showed a higher level of gcyA expression than the parental wildtype strain; it supports the hypothesis that GcyX negatively regulates transcription of gcyA. However, another mutant cgrA did not show a significant effect on gcyA expression but severely reduced gcyB expression. This latter result indicates that CgrA positively regulates the gcyX2-gcyB-gcyC-cgrA operon. Lastly, an additional flow cytometry assay was performed to see if cGMP synthesis deficient mutants did not show any disruptions of encystment. Cysts were produced by the wildtype strain as well as in the gcyA, and gcyX mutant strains. It indicates that cGMP production is not related to the cyst production in A. brasilense. These results collectively reveal important new information on the regulation of cGMP production by A. brasilense.

How long is Analysis of a Gene Cluster that is Responsible for CGMP Production by the Nitrogen Fixing Symbiont Azospirillum Brasilense?

Analysis of a Gene Cluster that is Responsible for CGMP Production by the Nitrogen Fixing Symbiont Azospirillum Brasilense by HyunAh Kim is 0 pages long, and a total of 0 words.

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