Exploring nodulation signaling in the rhizobium-legume symbiosis for improving existing and building synthetic symbioses
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Authors
Panchal, Aakanx
Date
2024-10-02
Type
thesis
Language
eng
Keyword
biology , rhizobium , legume
Alternative Title
Abstract
Sustainable alternatives to current methods of nitrogen supplementation in agricultural fields include improving existing symbioses and building synthetic symbioses. Legume root nodules are often occupied by native low N2-fixing rhizobia such that the benefit of inoculant high N2-fixing rhizobia is diminished. A strategy to potentially solve this problem would be building exclusive compatibility between inoculant rhizobia and target legumes. By engineering inoculant rhizobia to produce Nod factors that native rhizobia are not expected to produce and by engineering the legume to produce Nod factor receptors specific to those Nod factors, a simple version of exclusive compatibility could be implemented. To test the feasibility of this approach, I constructed a plasmid carrying the Nod factor receptor genes of Lotus japonicus for its eventual transformation into Medicago truncatula. I tested a transformation protocol for M. truncatula using an empty vector and observed that it was successful. I also worked towards constructing a Mesorhizobium japonicum strain that can only produce Sinorhizobium meliloti Nod factors, which requires deleting the M. japonicum Nod factor biosynthesis genes, by testing several methods for conjugating integrative plasmids capable of making gene deletions into M. japonicum. I could not obtain transconjugants of M. japonicum from integrative plasmids but I could obtain transconjugants carrying replicative plasmids. On the side of synthetic symbioses, I worked towards characterizing the S. meliloti transcription factor NodD1, which interacts with legume-exuded luteolin and transcriptionally activates Nod factor biosynthesis. Engineering the substrate-binding site of NodD1 to accept molecules exuded by non-legumes could be a way to build communication between rhizobia and non-legumes. Since the interaction of NodD1 and luteolin is not well characterized, there is also fundamental knowledge to be gained in studying NodD1, such as through X-ray crystallography. I worked towards this by constructing plasmids for and testing NodD1 overexpression. Once I observed NodD1 overexpression, I tested several methods to obtain soluble NodD1 but could not find any successful methods. I have set up alternative NodD1-overexpressing constructs for future solubility tests. Overall, some progress has been made in the difficult journey towards the sustainable growth of plants.
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ProQuest PhD and Master's Theses International Dissemination Agreement
Intellectual Property Guidelines at Queen's University
Copying and Preserving Your Thesis
This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner.
Attribution 4.0 International