Date of Award

4-2018

Degree Name

Master of Science

Department

Biological Sciences

First Advisor

Dr. Silvia Rossbach

Second Advisor

Todd Barkman, Ph.D.

Third Advisor

Jian Yao, Ph.D.

Keywords

Pinitol, d-chiro-inositol, nodules, sinorhizobium melitoti, nitrogen fixation

Access Setting

Masters Thesis-Open Access

Abstract

Sinorhizobium meliloti, which is the nitrogen fixing symbiont of Medicago sativa (alfalfa), was used to test the hypothesis that D-chiro-inositol is an intermediate of pinitol catabolism and that pinitol catabolism plays a role in S. melilot s ability to compete for nodule occupancy. An analytical method, involving gas chromatography and mass spectrometry (GCMS), was developed to detect and separate pinitol and D-chiro-inositol. The S. meliloti wild type, and three isogenic mutant strains (idhA, mocD, and mocE) were grown in minimal medium with pinitol and pinitol plus glycerol. The S. meliloti wild type can utilize pinitol as a sole carbon source, whereas the mutant strains cannot. The idhA mutant removed 50% of the detectable pinitol from the medium when grown with pinitol plus glycerol and showed accumulation of Dchiro- inositol. Therefore, it was concluded that D-chiro-inositol is indeed an intermediate of pinitol catabolism. To test the ability of pinitol catabolism mutants to compete with the wild type for nodule occupancy, alfalfa plants were inoculated with a 1:1 ratio of wild type S. meliloti and the idhA, mocD, and mocE mutants. Plants nodules were collected at eight and 16 weeks and the percentage of the nodule inhabitants was determined. Each mutant was found to be outcompeted by the wild type by week 16. Therefore, it was concluded that the ability to catabolize pinitol does indeed play a role in nodule occupancy.

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