Abstract
The Biolistic® microprojectile DNA-delivery method was used to test the usefulness in conifers of eight gene constructs based on the 35S promoter, the AMV translational enhancer, and gene fusion between the P-glucuronidase and the neomycin phosphotransferase II genes. The evaluation was done with embryogenic cells of Picea glauca, where the relative strengths of the promoters were 35S-35S-AMVE>35S-AMVE>35S-35S>35S as evaluated by transient gene expression. The fusion gene of GUS and NPT II gave lower levels of transient gene expression than the unfused GUS gene as detected by X-GLU histochemical assays. Experiments comparing the EM promoter of wheat and the 35S-35S-AMVE promoter (with and without fusion between GUS and NPT II) were done in Picea rubens, P. mariana, P. glauca, and Larix x eurolepis. The unfused gene with the 35S-35S-AMVE promoter gave higher levels of transient gene expression than the fused GUS-NPT II gene. The fluorescent MUG assay was more sensitive than the histochemical X-GLU assay to detect the activity of the β-glucuronidase gene.
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Abbreviations
- AMV:
-
alfalfa mosaic virus
- AMVE:
-
alfalfa mosaic virus translational enhancer
- EM:
-
protein of mature wheat embryo
- GUS:
-
P-glucuronidase gene
- MUG:
-
4-methylumbelliferyl β-D-glucuronide
- NPT II:
-
neomycin phosphotransferase
- X-GLU:
-
5-bromo-4-chloro-3-indolyl β-D-glucuronic acid
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Charest, P.J., Caléro, N., Lachance, D. et al. Microprojectile-DNA delivery in conifer species: factors affecting assessment of transient gene expression using the β-glucuronidase reporter gene. Plant Cell Reports 12, 189–193 (1993). https://doi.org/10.1007/BF00237051
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DOI: https://doi.org/10.1007/BF00237051