THE EFFECT OF ENVIRONMENTAL-CONDITIONS ON THE EXPRESSION OF DISEASE IN CULTURED-TISSUES OF BRASSICA-NAPUS SSP OLEIFERA INFECTED WITH LEPTOSPHAERIA-MACULANS

As a basis for devising an in vitro screening programme, culture condi tions were optimized so that tissue cultures from two resistant cultiv ars of Brassica napus ssp. oleifera (Mikado, Bienvenu) and two suscept ible cultivars (Lesira, Ceres) could be differentiated using a disease scoring scheme, when inoculated with Leptosphaeria maculans. Tissues inoculated included thin cell layer explants from soil-grown plants an d in vitro-grown shoot cultures and callus tissue formed on such expla nts. The period of incubation and the incubation temperature were of i mportance in the development of differential disease reactions. Increa sing temperature generally resulted in an increase in infection and to o great an incubation period resulted in total overgrowth of the tissu e. Increasing concentrations (1 X 10(-6) M-1 X 10(-4) M) of the auxins 1-naphtbylacetic acid (NAA), 2,4-dichlorophenoxyacetic acid (2,4-D) a nd indole-3-acetic acid (IAA) in the culture medium, resulted in a dec rease in disease score of the thin cell layer (TCL) explants from soil -grown plants. The cytokinins examined 6-benzylaminopurine (BAP) and 6 -4-hydroxy-3-methyl-2-enylaminopurine (zeatin), reduced the extent of infection of the TCL explants when used in combination with the auxin NAA. Medium containing NAA at a concentration of 1 x 10(-6) M in combi nation with BAP at a concentration of 1 x 10(-6) or 1 x 10(-4) M allow ed differentiation of the disease reactions of the resistant and susce ptible cultivars, when the explants were incubated for 10 days at 20-d egrees-C after inoculation. Similar conditions of incubation and the a ddition of NAA (1 x 10(-6) M) combined with BAP (1 x 10(-5) M) to the medium also allowed the differentiation of the disease reactions on TC L explants from stems of in vitro shoot cultures of the cultivars Mika do and Lesira. Increasing concentrations of the auxin NAA and the cyto kinin BAP resulted in a reduction in the mean disease score of the cal lus tissue produced on TCL explants from soil-grown plants, and NAA (1 x 10(-5) M) combined with BAP (1 x 10(-6) or 1 X 10(-5) M) allowed di fferentiation of resistance and susceptibility in callus tissues when incubated for 5 days at 20-degrees-C. 2,4-D did not allow differentiat ion of the cultivars. This was in contrast to the inoculation of callu s tissue attached to TCL explants of in vitro shoot cultures, where co mbinations of 2,4-D and BAP at concentrations of 1 x 10(-6) M allowed differentiation of the resistant and susceptible cultivars. These find ings provide a basis for designing selection protocols of value in bot h traditional as well as in vitro breeding programmes to select lines of oilseed rape with resistance/novel resistance to L. maculans.