Gene silencing using a heat-inducible RNAi system in Arabidopsis

Abstract

Controlling gene expression during plant development is an efficient tool to explore gene function. In this paper, we describe a gene expression system driven by a heath shock gene promoter (HSP18.2), to trigger the expression of an intron-containing inverted repeat. RNA interference became a powerful way for gene functional analysis by reverse genetic approaches. However, constitutive gene silencing cannot be used with genes involved in fundamental processes such as embryo viability. Inducible promoters provide an alternative approach for temporal and spatial gene expression control. And we described here a new, system, complementary to those using chemical gene inducers. To evaluate the efficiency of this system, RNA corresponding to the phytoene desaturase gene of Arabidopsis thaliana was used as a reporter gene in transgenic plants and a comparative study was performed using either the CaMV35S constitutive promoter or the HSP18.2 inducible promoter.

Controlled Expression of Recombinant Proteins in Physcomitrella patens by a Conditional Heat-shock Promoter: a Tool for Plant Research and Biotechnology

“…Thus, in Nicotiana tabacum the promoter hsp17.6L from soybean can control antibiotic resistance in a temperature-dependent manner (Severin and Schoffl, 1990) as well as the expression of the FLP recombinase in maize (Lyznik et al, 1995) and Arabidopsis thaliana (Kilby et al, 2000). Similarly, the A. thaliana hsp18.2 promoter was successfully used as a heat-inducible expression system in tobacco BY2 cells (Yoshida et al, 1995), and to achieve heat-inducible RNAi expression in A. thaliana (Masclaux et al, 2004). Recently, using the soybean Gmhsp17.3B promoter, inducible insertional mutagenesis, based on Ac-Ds transposons, was developed in A. thaliana (Nishal et al, 2005).…”

Two Cytochrome P450 Monooxygenases Catalyze Early Hydroxylation Steps in the Potato Steroid Glycoalkaloid Biosynthetic Pathway

“…A 372-bp fragment of PGA1 cDNA that did not match over consecutive 10 nucleotides with PGA2 was PCR amplified using primer set U724/U725. The fragments were inserted in opposite orientations with the intron of the Arabidopsis (Arabidopsis thaliana) PDS gene (Masclaux et al, 2004) by XbaI/SacI digestion into the pKT19 vector, substituting for the GUS gene, under the control of the 35S promoter in the T-DNA region to produce the RNAi binary vector pKT226 (Supplemental Fig. S1).…”

T-DNA insertional mutagenesis in Arabidopsis: a tool for functional genomics

“…However, it is a laborious approach and results are often inconclusive or impossible to interpret (Azpiroz-Leehan and Feldmann, 1997). Recent reports of successful use of double stranded RNA mediated interference (RNAi) approach in C. elegans (Ashrafi et al 2003;Kamath and Ahringer, 2003), Arabidopsis (Wang and Waterhouse, 2001;Masclaux et al 2004) is certain to open new vistas for sequence-specific inhibition of gene function in Arabidopsis (Tuschl, 2003).…”