Induction and purification of chitinase in Brassica napus L. ssp. oleifera infected with Phoma lingam.

A pathogen-induced chitinase (EC 3.2.1.14) was isolated from cotyledons of oilseed rape (Brassica napus cv. Bienvenu) 8 d after inoculation with Phoma lingam. The purified chitinase has a molecular weight of 30 kDa, and an isoelectric point of approx. 9.1. A partial amino-acid sequence obtained after tryptic digestion of the protein shows high sequence similarity to basic chitinases from bean, tobacco, potato, Arabidopsis, barley and rice, as well as to acidic chitinases from tobacco and petunia. A close serological relationship was found between the chitinase isoenzyme and an isoenzyme from sugar-beet (Beta vulgaris L.). When resistant and susceptible cultivars were inoculated with P. lingam there was a significant difference in the increase in chitinase activity during the early stage after inoculation. The resistant cultivars showed a rapid increase in chitinase activity, in contrast to susceptible cultivars where an increase in activity was delayed until 24 h after infection. By measuring the chitinase activity from the mycelium of P. lingam, it was concluded that the increase in chitinase activity found in infected plants was of plant origin. The chitinase activity was found to be restricted to the site of pathogen attack and was not systemically induced in other parts of the plant.