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Partial characterization of a putative transcriptional regulator of the 4CL1 gene Jones, Jennifer Anne
Abstract
This thesis is intended to further our understanding of the mechanisms by which expression of phenylpropanoid genes, specifically parsley 4-coumarate : CoA ligase (4CL1), are developmental^ regulated. The 4CL enzyme is the third and final step in the core phenylpropanoid metabolic pathway. 4CL1-GUS fusions are specifically expressed in the xylem and floral organs in transgenic tobacco. In-vivo footprinting and sitedirected mutagenesis revealed a region spanning -78 to -120, known as footprint 56, of the 4CL1 promoter which is critical for 4CL expression. DNasel footprinting and gel shift assays using tobacco nuclear extracts indicated that a nuclear factor or complex of factors bind to this cis-element on the 4CL1 promoter in vitro. A putative transcription factor which binds to this footprint 56, termed 56BF, was isolated by using a footprint 56 oligonucleotide as a probe to screen a tobacco cDNA expression library. To test whether 56BF is involved in regulating expression of the 4CL1 gene, transgenic tobacco plants that over-expressed or were suppressed for its' accumulation were generated. 56BF recombinant protein was expressed in E. Coli to test the DNA-binding specificity of the protein. Results discussed in this thesis strongly suggest that 56BF does not regulate the 4CL1 gene in vivo.
Item Metadata
Title |
Partial characterization of a putative transcriptional regulator of the 4CL1 gene
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Creator | |
Publisher |
University of British Columbia
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Date Issued |
1998
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Description |
This thesis is intended to further our understanding of the mechanisms by which
expression of phenylpropanoid genes, specifically parsley 4-coumarate : CoA ligase
(4CL1), are developmental^ regulated. The 4CL enzyme is the third and final step in the
core phenylpropanoid metabolic pathway. 4CL1-GUS fusions are specifically expressed
in the xylem and floral organs in transgenic tobacco. In-vivo footprinting and sitedirected
mutagenesis revealed a region spanning -78 to -120, known as footprint 56, of
the 4CL1 promoter which is critical for 4CL expression. DNasel footprinting and gel
shift assays using tobacco nuclear extracts indicated that a nuclear factor or complex of
factors bind to this cis-element on the 4CL1 promoter in vitro. A putative transcription
factor which binds to this footprint 56, termed 56BF, was isolated by using a footprint 56
oligonucleotide as a probe to screen a tobacco cDNA expression library. To test whether
56BF is involved in regulating expression of the 4CL1 gene, transgenic tobacco plants
that over-expressed or were suppressed for its' accumulation were generated. 56BF
recombinant protein was expressed in E. Coli to test the DNA-binding specificity of the
protein. Results discussed in this thesis strongly suggest that 56BF does not regulate the
4CL1 gene in vivo.
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Extent |
5221969 bytes
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Genre | |
Type | |
File Format |
application/pdf
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Language |
eng
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Date Available |
2009-05-25
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Provider |
Vancouver : University of British Columbia Library
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Rights |
For non-commercial purposes only, such as research, private study and education. Additional conditions apply, see Terms of Use https://open.library.ubc.ca/terms_of_use.
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DOI |
10.14288/1.0088487
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URI | |
Degree | |
Program | |
Affiliation | |
Degree Grantor |
University of British Columbia
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Graduation Date |
1998-11
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Campus | |
Scholarly Level |
Graduate
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Aggregated Source Repository |
DSpace
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Rights
For non-commercial purposes only, such as research, private study and education. Additional conditions apply, see Terms of Use https://open.library.ubc.ca/terms_of_use.