IDENTIFICATION AND CHARACTERIZATION OF cDNA INDUCED BY LOW TEMPERATURE IN SNOW MOULD-RESISTANT AND FREEZING-TOLERANT WINTER WHEAT
D GAUDET, M FRICK, A LAROCHE, R HUEL, P AUDY and J DAVERON
Research Centre, Agriculture and Agri-Food Canada, PO Box 3000, Lethbridge, Alberta, Canada T1J 4B1
Background and objectives
In the Parkland area of Canada, high levels of snow mould resistance and freezing tolerance are necessary for winter wheat survival. Canadian winter wheat cultivars are freezing-tolerant but susceptible to snow mould. Snow mould-resistant cultivars are too tender to survive in western Canada. Current protocols for screening for both snow mould resistance and freezing tolerance are time-consuming and often produce inconsistent results. In wheat, snow mould resistance is governed by the additive action of two or three genes whereas many genes govern the expression of freezing tolerance. Both forms of resistance are induced by acclimation at low temperature. The objectives of this project are to understand the mechanism(s) of snow mould resistance and apply molecular techniques to the development snow mould-resistant winter wheats. We report the isolation and characterization of cDNAs that are related to snow mould resistance.
Material and methods
The snow mould-resistant, freezing-tolerant doubled haploid winter wheat line DH++, was generated from the F1 of a cross between Norstar, a freezing tolerant but snow mold susceptible cultivar, and PI181268, a snow moud-resistant but tender cultivar. RNA was isolated from the crowns of seedlings grown for 4 weeks at 2°C and used to construct a cDNA subtraction library . Low temperature-induced genes were isolated from the library and their expression characterized in snow mould-resistant and susceptible winter wheat genotypes. Sequences related to either freezing tolerance and snow mould resistance were identified by Northern analyses and RT-PCR. Putative identity of cDNAs was verified using sequence homology to non-redundant databases .
Results and conclusions
Eight different gene products induced at low temperature, appeared to be related to snow mould resistance. The differential expression of these gene products in cultivars differing in level of snow mould resistance and freezing tolerance strongly suggest that they are related to snow mould resistance. These novel clones do not show significant homology to sequences present in the different databases. The expression of the sequences will be discussed in relation to low temperature induction, tissue specificity, and repression under dehardening conditions.
1. Patel M, Sive H, 1996. In Current Protocols in Molecular Biology (Ausubel FM et al., eds). Wiley, New York, pp. 5.9.1-5.9.20.
2. Altschul SF, Gish W, Miller W, Myers EW, Lipman DJ, 1990. Journal of Molecular Biology 215, 403-410