Abstract
We describe a new luciferase reporter gene,luc INT, for early detection of luciferase activity inAgrobacterium transformation studies, and present improved techniques for the extraction of luciferase that decrease the time needed to quantitate luciferase activity. Theluc INT reporter gene combines the PIV2 intron fromGUS INT withluc *, the modified luciferase gene.luc INTis expressed in plant cells but not inAgrobacterium, allowing earlier detection of gene expression in the presence ofAgrobacterium during transformations in tobacco leaf discs. Stable expression levels ofluc INT andluc * in tobacco suspension cultures are compared for two different promoters.
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Abbreviations
- 35S:
-
35S promoter of cauliflower mosaic virus
- CaMV:
-
cauliflower mosaic virus
- CLR:
-
cell-culture lysis reagent
- GFP:
-
green fluorescent protein
- GUS:
-
β-glucuronidase
- GUSINT :
-
uidA with the PIV2 intron
- luc :
-
cDNA encoding luciferase
- luc * :
-
a specially modifiedluc
- lucINT :
-
luc cDNA with the PIV2 intron
- LAR:
-
luciferin assay reagent
- nosT :
-
gene encoding nopaline
- ocs :
-
gene encoding octopine synthase promoter region
- PCC:
-
photon-counting camera
- RT-PCR:
-
reverse transcription-PCR
- uidA :
-
gene encoding β-glucuronidase
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The nucleotide sequence data for the gene will appear in the EMBL, GenBank and DDBJ Nucleotide Sequence Databases under the accession number U84006.
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Luke Mankin, S., Allen, G.C. & Thompson, W.F. Introduction of a plant intron into the luciferase gene ofPhotinus pyralis . Plant Mol Biol Rep 15, 186–196 (1997). https://doi.org/10.1007/BF02812270
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DOI: https://doi.org/10.1007/BF02812270