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Title: A molecular toolkit for high-throughput identification of competitive and effective rhizobial strains
Author: Suarez, Marcela Mendoza
ISNI:       0000 0004 7966 2014
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 2018
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Legumes tend to become nodulated by highly competitive but not necessarily effective rhizobial strains, resulting in less optimal plant growth. Research focused on the identification of both competitive and highly effective nitrogen-fixing strains has been limited because, so far, the only way to test competitive rhizobial strains has been to isolate individual strains and compare them one at a time in large-scale plant growth assays. We have developed a library of novel reporter plasmids (Plasmid IDs) compatible with Golden Gate cloning which consist of i) a promoter-probe vector (pOGG026) suitable for studies where antibiotic selection is not possible, ii) the maker gene sfGFP under the control of a synthetic consensus promoter (psNifH) to drive nodule-specific expression in biovars and strains of Rhizobium leguminosarum and which can be used as a proxy for nitrogenase activity assays, and iii) a 12-nucleotide specific sequence to mark R. leguminosarum strains. Additionally, we developed reporter plasmids that express biomarkers, such as gusA, celB and mCherry suitable for rhizobial screening in the rhizosphere. The plasmid pOGG026 allowed us to develop a new plasmid-based system to mark Rhizobium strains with GusA/CelB and evaluate their competitiveness. Results demonstrated the proof of concept of Plasmid IDs as a tool for high-throughput identification of competitive and effective rhizobial strains. The application of this tool in a competition assay among eighty-four Tagged-strains, in non-sterilised agricultural soil containing indigenous rhizobia, allowed us to identify an effective and super competitive strain, in addition to mixed nodules occupied by up to four Tagged-strains. This tool has the potential to revolutionise the search for elite indigenous rhizobia, by reducing the costs involved in identifying competitive and effective rhizobial strains.
Supervisor: Poole, Philip Sponsor: CONACYT
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Plant-bacteria interactions