Modulation of root antioxidant status to delay cassava post-harvest physiological deterioration

Michael Page

Research output: ThesisDoctoral Thesis

Abstract

Cassava ranks seventh in terms of worldwide crop production, providing a staple for over half a billion people. The production of cassava is limited by several factors, with post-harvest physiological deterioration (PPD) of storage roots a major constraint. PPD is a process initiated on harvesting and mediated by reactive oxygen species (ROS) that ultimately renders storage roots unpalatable and unmarketable. It is similar to a conventional plant wound response, but crucially lacks efficient wound repair and down-regulation of stress signalling. Therefore, the strategy utilised here to modulate PPD focussed on increasing the ROS scavenging ability of storage root tissue through a biotechnological approach.

Three expression plasmids were produced, harbouring cassava genes encoding the antioxidant enzymes APX, CAT and SOD under the control of the storage rootspecific StPAT promoter. In addition, a reporter expression plasmid was created, with StPAT driving the expression of GusP. Transgenic Arabidopsis plants containing the StPAT::GusP cassette demonstrated root-specific GusP staining. Non-root tissue also showed wound-inducible GusP activity conferred by the StPAT promoter. This novel activity was detected almost immediately after wounding and occurred independently of ethylene, MeJa and ROS. The 3’ 261 bp of the StPAT promoter was sufficient to confer wound-inducible expression and contained putative wound responsive cis regulatory motifs. Analysis of PATATIN function indicated a role during early responses to wounding in the liberation of free fatty acids from cell membranes.

Over-expression of the target genes in the model plant Arabidopsis increased the antioxidant enzyme activity in the roots of selected lines. Transgenic plants generally exhibited similar levels of oxidative stress resistance to wild-type plants, a result due in part to the efficient nature of the oxidative stress response of Arabidopsis – the APX activity of wild-type plants increased to transgenic levels under H2O2 stress. However, PPD in cassava is at least partially the result of a poor antioxidant response to harvesting, and so transformation of cassava with the expression plasmids remained a viable strategy. Transgenic cassava plants harbouring the expression cassettes are being generated and will soon be assessed for PPD resistance.

LanguageEnglish
QualificationPh.D.
Awarding Institution
  • University of Bath
Supervisors/Advisors
  • Beeching, John, Supervisor
  • Hooley, Richard, Supervisor
Award date1 May 2009
StatusUnpublished - May 2009

Fingerprint

cassava
plant damage
deterioration
antioxidants
genetically modified organisms
plasmids
promoter regions
oxidative stress
Arabidopsis
stress tolerance
crop production
cell membranes
ethylene
stress response
superoxide dismutase
genes
enzyme activity
fatty acids
oxygen
enzymes

Keywords

  • cassava
  • reactive oxygen species
  • antioxidant
  • deterioration

Cite this

Modulation of root antioxidant status to delay cassava post-harvest physiological deterioration. / Page, Michael.

2009. 203 p.

Research output: ThesisDoctoral Thesis

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AB - Cassava ranks seventh in terms of worldwide crop production, providing a staple for over half a billion people. The production of cassava is limited by several factors, with post-harvest physiological deterioration (PPD) of storage roots a major constraint. PPD is a process initiated on harvesting and mediated by reactive oxygen species (ROS) that ultimately renders storage roots unpalatable and unmarketable. It is similar to a conventional plant wound response, but crucially lacks efficient wound repair and down-regulation of stress signalling. Therefore, the strategy utilised here to modulate PPD focussed on increasing the ROS scavenging ability of storage root tissue through a biotechnological approach. Three expression plasmids were produced, harbouring cassava genes encoding the antioxidant enzymes APX, CAT and SOD under the control of the storage rootspecific StPAT promoter. In addition, a reporter expression plasmid was created, with StPAT driving the expression of GusP. Transgenic Arabidopsis plants containing the StPAT::GusP cassette demonstrated root-specific GusP staining. Non-root tissue also showed wound-inducible GusP activity conferred by the StPAT promoter. This novel activity was detected almost immediately after wounding and occurred independently of ethylene, MeJa and ROS. The 3’ 261 bp of the StPAT promoter was sufficient to confer wound-inducible expression and contained putative wound responsive cis regulatory motifs. Analysis of PATATIN function indicated a role during early responses to wounding in the liberation of free fatty acids from cell membranes. Over-expression of the target genes in the model plant Arabidopsis increased the antioxidant enzyme activity in the roots of selected lines. Transgenic plants generally exhibited similar levels of oxidative stress resistance to wild-type plants, a result due in part to the efficient nature of the oxidative stress response of Arabidopsis – the APX activity of wild-type plants increased to transgenic levels under H2O2 stress. However, PPD in cassava is at least partially the result of a poor antioxidant response to harvesting, and so transformation of cassava with the expression plasmids remained a viable strategy. Transgenic cassava plants harbouring the expression cassettes are being generated and will soon be assessed for PPD resistance.

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