Characterization of phenylalanine ammonia-lyase (PAL) gene in wounded lettuce tissue
AssigneeThe Regents of the University of California
InventorsMikal E. Saltveit, Reinaldo Campos, Hiroyuki Nonogaki, Trevor Suslow
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Abstract
The invention provides phenylalanine ammonia-lyase (PAL) sequences, and constructs and methods related thereto. In one aspect, Lactuca sativa PAL sequences are isolated and characterized. The invention also provides novel sequences associated with PAL polynucleotide sequences and methods using recombinant techniques to express proteins from such sequences. In addition, methods are provided for using such sequences to transform host cells and alter wound induced gene activity or expression.
Claims23
An isolated nucleic acid comprising a nucleotide sequence or a fragment thereof encoding the amino acid sequence set forth in SEQ ID NO:1 or SEQ ID NO:3.
A nucleic acid fragment that hybridizes to SEQ ID NO:2 or SEQ ID NO:4 under stringent hybridization conditions and has other than a nucleotide sequence as shown in FIG. 2.
An antibody that binds specifically to the amino acid sequence or portion thereof set forth in SEQ ID NO:1 or SEQ ID NO:3.
An isolated nucleic acid construct comprising a transcriptional initiation sequence operably linked to SEQ NO:2 or SEQ NO:4.
A degenerate primer pair based on Phenylalanine ammonia-lyase homologous sequences in closely related plants, wherein first primer of said paired primers is GAYCCNYTNAAYTGGGG and second primer of said paired primers is CCYTGPAARTTNCCNCCRTG.
A method of producing a transgenic cell having altered phenylalanine ammonia-lyase levels, said method comprising:
- introducing an expression cassette comprising a transcription initiation sequence operably linked to an open reading frame coding for SEQ ID NO:1 or SEQ ID NO:3 or an enzymatically active fragment thereof, and;
- growing said cell whereby said open reading frame is expressed and a cell having altered phenylalanine ammonia-lyase is produced.
A method for measuring the relative amount of phenylalanine ammonia-lyase levels in a tissue, said method comprising:
- contacting said tissue with antibodies specific for the amino acid sequence set forth in SEQ ID NO:1 or SEQ ID NO:3, and;
- comprising a detectable label wherein a change in the intensity of said detectable label in said tissue as compared to a control tissue is indicative of an increase or decrease of phenylalanine ammonia-lyase in said tissue.
The method of claim 19, wherein said antibodies are polyclonal.
The method of claim 20, wherein said antibodies are monoclonal.
A kit for measuring phenylalanine ammonia-lyase protein levels in an article of produce, comprising:
- antibodies specific for the amino acid sequence set forth in SEQ ID NO:1 or SEQ ID NO:3 or a particle thereof.
Antibodies specific for the amino acid sequence set forth in SEQ ID NO:1 or SEQ ID NO:3 or a particle thereof, wherein said antibodies are polyclonal.
Antibodies specific for the amino acid sequence set forth in SEQ ID NO:1 or SEQ ID NO:3 or a particle thereof, wherein said antibodies are monoclonal.
Dependencies are inferred from the claim text.