Characterization of a major cluster of genes involved in nitrogen fixation and another required for indole-3-acetic acid biosynthesis in the sugarcane endophyte, Acetobacter diazotrophicus

Persistent Link:
http://hdl.handle.net/10150/279953
Title:
Characterization of a major cluster of genes involved in nitrogen fixation and another required for indole-3-acetic acid biosynthesis in the sugarcane endophyte, Acetobacter diazotrophicus
Author:
Lee, Sunhee
Issue Date:
2001
Publisher:
The University of Arizona.
Rights:
Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
Abstract:
Acetobacter diazotrophicus is a true endophyte of sugarcane and is often found in plants grown in agricultural areas of low nitrogen fertilizer input. Results from our laboratory, using mutant strains of A. diazotrophicus unable to fix nitrogen, have shown that there are two beneficial effects of A. diazotrophicus on sugarcane: one dependent on nitrogen fixation, and the other independent of nitrogen fixation. A plant growth promoting substance like indole-3-acetic acid (IAA) may represent the latter effect that accounts for improved plant growth. My first project was to characterize the genes responsible for nitrogen fixation, and determine their regulation. In summary, I have isolated, sequenced, and analyzed the major 31.5 kb nif gene cluster, including both nif and associated genes of A. diazotrophicus. This cluster of 33 genes represents the largest and most complete assembly of contiguous nif/fix and associated genes characterized in any diazotrophic bacterial species. My second project has been to determine whether nitrogen fixation and/or IAA production are important for the ability of A. diazotrophicus to stimulate plant growth. In order to determine the role of IAA directly, mutants of A. diazotrophicus producing reduced amounts of IAA were generated by Tn5 mutagenesis. Among IAA - candidates, one excreting less than 6% of IAA compared to the parent strain was further characterized. The mutation was mapped to genes involved in cytochrome c biogenesis (ccm genes-c&barbelow;ytochrome c&barbelow; m&barbelow;aturation genes). A Nif -/Iaa- double mutant and Nif- mutant were constructed by inserting a chloramphenicol cassette into nifD region. Plant inoculation experiments using mutant strains also demonstrated that A. diazotrophicus could stimulate plant growth regardless of N availability, as evidenced by the significant growth difference between plants inoculated with wild type and uninoculated plants. Under N-limiting conditions plants inoculated with wild type had greater height and biomass than plants inoculated with Nif- or Nif -/Iaa- mutants, suggesting nitrogen fixation by A. diazotrophicus stimulates sugarcane growth. Plants inoculated with Iaa- mutants were always comparable to uninoculated plants regardless of N availability, indicating that IAA biosynthesis is a major bacterial factor influencing sugarcane growth, particularly under N-sufficient conditions.
Type:
text; Dissertation-Reproduction (electronic)
Keywords:
Biology, Molecular.; Biology, Microbiology.; Agriculture, Plant Pathology.
Degree Name:
Ph.D.
Degree Level:
doctoral
Degree Program:
Graduate College; Plant Diseases
Degree Grantor:
University of Arizona
Advisor:
Kennedy, Christina

Full metadata record

DC FieldValue Language
dc.language.isoen_USen_US
dc.titleCharacterization of a major cluster of genes involved in nitrogen fixation and another required for indole-3-acetic acid biosynthesis in the sugarcane endophyte, Acetobacter diazotrophicusen_US
dc.creatorLee, Sunheeen_US
dc.contributor.authorLee, Sunheeen_US
dc.date.issued2001en_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.description.abstractAcetobacter diazotrophicus is a true endophyte of sugarcane and is often found in plants grown in agricultural areas of low nitrogen fertilizer input. Results from our laboratory, using mutant strains of A. diazotrophicus unable to fix nitrogen, have shown that there are two beneficial effects of A. diazotrophicus on sugarcane: one dependent on nitrogen fixation, and the other independent of nitrogen fixation. A plant growth promoting substance like indole-3-acetic acid (IAA) may represent the latter effect that accounts for improved plant growth. My first project was to characterize the genes responsible for nitrogen fixation, and determine their regulation. In summary, I have isolated, sequenced, and analyzed the major 31.5 kb nif gene cluster, including both nif and associated genes of A. diazotrophicus. This cluster of 33 genes represents the largest and most complete assembly of contiguous nif/fix and associated genes characterized in any diazotrophic bacterial species. My second project has been to determine whether nitrogen fixation and/or IAA production are important for the ability of A. diazotrophicus to stimulate plant growth. In order to determine the role of IAA directly, mutants of A. diazotrophicus producing reduced amounts of IAA were generated by Tn5 mutagenesis. Among IAA - candidates, one excreting less than 6% of IAA compared to the parent strain was further characterized. The mutation was mapped to genes involved in cytochrome c biogenesis (ccm genes-c&barbelow;ytochrome c&barbelow; m&barbelow;aturation genes). A Nif -/Iaa- double mutant and Nif- mutant were constructed by inserting a chloramphenicol cassette into nifD region. Plant inoculation experiments using mutant strains also demonstrated that A. diazotrophicus could stimulate plant growth regardless of N availability, as evidenced by the significant growth difference between plants inoculated with wild type and uninoculated plants. Under N-limiting conditions plants inoculated with wild type had greater height and biomass than plants inoculated with Nif- or Nif -/Iaa- mutants, suggesting nitrogen fixation by A. diazotrophicus stimulates sugarcane growth. Plants inoculated with Iaa- mutants were always comparable to uninoculated plants regardless of N availability, indicating that IAA biosynthesis is a major bacterial factor influencing sugarcane growth, particularly under N-sufficient conditions.en_US
dc.typetexten_US
dc.typeDissertation-Reproduction (electronic)en_US
dc.subjectBiology, Molecular.en_US
dc.subjectBiology, Microbiology.en_US
dc.subjectAgriculture, Plant Pathology.en_US
thesis.degree.namePh.D.en_US
thesis.degree.leveldoctoralen_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.disciplinePlant Diseasesen_US
thesis.degree.grantorUniversity of Arizonaen_US
dc.contributor.advisorKennedy, Christinaen_US
dc.identifier.proquest3002543en_US
dc.identifier.bibrecord.b41434286en_US
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