A multisite gateway-based toolkit for targeted gene expression and hairpin RNA silencing in tomato fruits
Article first published online: 18 FEB 2009
© 2009 The Authors. Journal compilation © 2009 Blackwell Publishing Ltd
Plant Biotechnology Journal
Volume 7, Issue 3, pages 298–309, April 2009
How to Cite
Estornell, L. H., Orzáez, D., López-Peña, L., Pineda, B., Antón, M. T., Moreno, V. and Granell, A. (2009), A multisite gateway-based toolkit for targeted gene expression and hairpin RNA silencing in tomato fruits. Plant Biotechnology Journal, 7: 298–309. doi: 10.1111/j.1467-7652.2009.00402.x
- Issue published online: 8 MAR 2009
- Article first published online: 18 FEB 2009
- Received 8 October 2008; revised 16 December 2008; accepted 18 December 2008.
- fruit promoter;
- hairpin RNA;
- recombinant protein;
A collection of fruit promoters, reporter genes and protein tags has been constructed in a triple-gateway format, a recombination-based cloning system that facilitates the tandem assembly of three DNA fragments into plant expression vectors. The new pENFRUIT collection includes, among others, the classical tomato-ripening promoters E8 and 2A11 and a set of six new tomato promoters. The new promoter activities were characterized in both transient assays and stable transgenic plants. The range of expression of the new promoters comprises strong (PNH, PLI), medium (PLE, PFF, PHD) and weak (PSN) promoters driving gene expression preferentially in the fruit, and covering a wide range of tissues and developmental stages. Together, a total of 78 possible combinations for the expression of a gene of interest in the fruit, plus a set of five reporters for new promoter analysis, was made available in the current collection. Moreover, the pENFRUIT promoter collection is adaptable to hairpin RNA strategies aimed at tissue/organ-specific gene silencing with only an additional cloning step. The pENFRUIT toolkit broadens the spectrum of promoter activities available for fruit biotechnology and fundamental research, and bypasses technical difficulties of current ligase-dependent cloning techniques in the construction of fruit expression cassettes. The pENFRUIT vector collection is available for the research community in a plasmid repository, facilitating its accessibility.