dc.contributor.advisor |
MacRae, Elspeth |
en |
dc.contributor.advisor |
Gardner, Richard |
en |
dc.contributor.author |
Fung, Raymond W.M. |
en |
dc.date.accessioned |
2007-08-06T05:23:36Z |
en |
dc.date.available |
2007-08-06T05:23:36Z |
en |
dc.date.issued |
2003 |
en |
dc.identifier |
THESIS 06-221 |
en |
dc.identifier.citation |
Thesis (PhD--Biological Sciences)--University of Auckland, 2003 |
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dc.identifier.uri |
http://hdl.handle.net/2292/1265 |
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dc.description |
Full text is available to authenticated members of The University of Auckland only. |
en |
dc.description.abstract |
Sucrose-phosphate synthase (SPS) is the key sucrose-synthesizing enzyme in plants. All publicly available SPS gene sequences can be classified phylogenetically into three families, designated A, B and C. Members of the same family from different species were found to have similar intron/exon genomic organization. One family B gene was cloned from kiwifruit (Actinidia chinensis) and shown to be a single gene family.
Three cDNA clones were also characterized from SPS gene family A in kiwifruit. The clones (designated A1, A2 and A3) are closely related, sharing 94-96% nucleotide identity in the coding region, and ca 67% nucleotide identity to Family B. RT-PCR was used to monitor transcripts of the four kiwifruit genes, using primers in the gene-specific 3' untranslated regions. All three family A genes were expressed in all tissues, but only A1 was present at high abundance and responded to developmental and environmental stimuli. The A1 transcript increased in fruit in response to treatment with exogenous ethylene and again during the ripening climacteric, but was not affected by low temperature. A1 expression was also high in senescent leaves, stems and in flower buds. In contrast, transcript levels of A2 and A3 are expressed constitutively. The B-family transcript was detectable in roots, albeit at very low levels and in all other tissues examined. It was increased in low temperature and in fruit at the climacteric (but not with applied ethylene). This is the first demonstration of differential expression patterns for highly homologous SPS genes in any plant species. The high complexity of these SPS gene expression patterns help to explain the lack of correlation of previous transcript analysis with SPS enzymatic activity found in kiwifruit and in other plants.
One Family A member, SPS-A2, encoded a longer predicted open reading frame than the other two A-family genes, with a unique C-terminal extension of 102 amino acids. Anti-SPSA2 antibody was raised against a peptide synthesized from this C-terminal region. Initial Western blots of total protein extracted from kiwifruit fruit revealed a single 120-kDa immuno-reactive polypeptide. However, attempts to determine the specificity of the antibody by expression of SPS-A2 protein fragments in Escherichia coli were unsuccessful.
The nucleotide sequence of the promoter region of the SPS-A2 gene contained several putative response elements for environmental and developmental triggers like light, sucrose, gibberellin acid, ethylene, low temperature and abscisic acid. Four deleted derivatives of this promoter were constructed upstream of the luciferase reporter gene. However, no expression was detected in a transient expression system following delivery into apple protoplasts. Further A2 promoter analysis was performed stable transgenic plants of Arabidopsis GUS expression was detected specifically in the vascular tissues and in young seedlings. |
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dc.language.iso |
en |
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dc.publisher |
ResearchSpace@Auckland |
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dc.relation.ispartof |
PhD Thesis - University of Auckland |
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dc.relation.isreferencedby |
UoA99158722014002091 |
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dc.rights |
Restricted Item. Available to authenticated members of The University of Auckland. |
en |
dc.rights.uri |
https://researchspace.auckland.ac.nz/docs/uoa-docs/rights.htm |
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dc.title |
A multigene family encoding sucrose phosphate synthase in kiwifruit |
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dc.type |
Thesis |
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thesis.degree.discipline |
Biological Sciences |
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thesis.degree.grantor |
The University of Auckland |
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thesis.degree.level |
Doctoral |
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thesis.degree.name |
PhD |
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dc.rights.holder |
Copyright: The author |
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pubs.rights |
Items in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated. |
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dc.identifier.wikidata |
Q112858513 |
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