1.4.1
ISOLATION AND CHARACTERIZATION OF A GENE EXPRESSED IN SALIX VIMINALIS INOCULATED WITH MELAMPSORA EPITEA

L JOHANSSON1 and P SAARIKOSKI2

1Department of Plant Pathology, Unit 1, and 2Department of Entomology, Swedish University of Agricultural Sciences, Box 7044, S-750 07 Uppsala, Sweden

Background and objectives
Salix is an important crop in Sweden. As with all crops cultivated intensively, Salix is exposed to epidemics. Its most important disease presently is willow leaf rust caused by Melampsora epitea. Rust infections can severely affect growth, reducing biomass production by up to 40% in short-rotation forests. Although a lot of work has been done on interactions and resistance in the Salix-Melampsora pathosystem, there are only a few reports available concerning the molecular response in Salix to pathogenic attack. To better understand the molecular mechanisms of resistance in Salix, we are searching for genes involved in resistance mechanisms.

Materials and methods
From one clone of S. viminalis (78183), two single-spore isolates were identified as two different pathotypes (86VIM and 77DAS) of M. epitea, based on their pathogenicity on different willow clones. Leaves from plants inoculated with the different rust pathotypes were collected daily, from the day of inoculation to day 10. RNA was extracted from these leaves for analysis by the differential mRNA display technique (DDRT-PCR). DNA fragments of interest were cloned into a TA vector and sequenced.

Results and conclusions
We have cloned and sequenced a 520-bp fragment of a putative receptor-like protein kinase gene. This fragment is expressed when S. viminalis is inoculated with rust isolated from S. dasyclados (77DAS), but is not expressed when inoculated with rust from S. viminalis (86VIM). This fragment shows a high degree of homology with the kinase domain of two receptor-like protein kinases from Arabidopsis thaliana. The role of the receptor-like protein kinases in a plant's defence against pathogens is not clear. However, there seems to be a strong relationship between protein kinases and the oxidative burst in plant defence [1]. Some of the receptor-like protein kinases are presumed to be candidates for disease resistance genes in plants . In Brassica the receptor-like protein kinases are known to act in the self-incompatibility response [2]. In this case, one theory is that the receptor-like protein kinase might form a signalling complex with products from resistance genes [2]. Further sequencing and characterization of this protein kinase gene is in progress.

References
1. Low PS, Merida JR, 1996. Physiologia Plantarum 96, 533-542.
2. Braun DM, Walker JC, 1996. Trends in Biochemical Science 21, 70-73.