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Plant Methods. 2010 Mar 11;6:8. doi: 10.1186/1746-4811-6-8.

Rapid expression of transgenes driven by seed-specific constructs in leaf tissue: DHA production.

Author information

1
CSIRO Food Futures National Research Flagship, PO Box 1600, Canberra, ACT 2601, Australia.
2
CSIRO Plant Industry, PO Box 1600, Canberra, ACT 2601, Australia.
#
Contributed equally

Abstract

BACKGROUND:

Metabolic engineering of seed biosynthetic pathways to diversify and improve crop product quality is a highly active research area. The validation of genes driven by seed-specific promoters is time-consuming since the transformed plants must be grown to maturity before the gene function can be analysed.

RESULTS:

In this study we demonstrate that genes driven by seed-specific promoters contained within complex constructs can be transiently-expressed in the Nicotiana benthamiana leaf-assay system by co-infiltrating the Arabidopsis thaliana LEAFY COTYLEDON2 (LEC2) gene. A real-world case study is described in which we first assembled an efficient transgenic DHA synthesis pathway using a traditional N. benthamiana Cauliflower Mosaic Virus (CaMV) 35S-driven leaf assay before using the LEC2-extended assay to rapidly validate a complex seed-specific construct containing the same genes before stable transformation in Arabidopsis.

CONCLUSIONS:

The LEC2-extended N. benthamiana assay allows the transient activation of seed-specific promoters in leaf tissue. In this study we have used the assay as a rapid preliminary screen of a complex seed-specific transgenic construct prior to stable transformation, a feature that will become increasingly useful as genetic engineering moves from the manipulation of single genes to the engineering of complex pathways. We propose that the assay will prove useful for other applications wherein rapid expression of transgenes driven by seed-specific constructs in leaf tissue are sought.

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