Microprojectile-DNA delivery in conifer species: factors affecting assessment of transient gene expression using the β-glucuronidase reporter gene
✍ Scribed by Pierre J. Charest; Nathalie Caléro; Denis Lachance; Raju S. S. Datla; Luc C. Duchêsne; Ed W. T. Tsang
- Publisher
- Springer
- Year
- 1993
- Tongue
- English
- Weight
- 855 KB
- Volume
- 12
- Category
- Article
- ISSN
- 0721-7714
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✦ Synopsis
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.