The role of birch protease-inhibitors and chitinases in resistance mechanisms against insects and diseases, respectively, was studied.
Chitinase. The native chitinases of silver birch (Betula pendula) were studied with isoelectric focusing. Several chitinases were found, but no variation in the chitinase pattern during the summer were noticed. Chimeric gene 4x35SCaMV - Chitinase4 (from sugar beet) attached to reporter gene 35S CaMV - uidA was transferred to birch using Agrobacterium. Those shoot lines expressing strong GUS activity are tested with western analysis and the best lines will be used in resistance tests.
Protease inhibitor. Trypsin inhibitors (TI) were partly purified from the leaves of wild birch. TI level was highest in early spring and stayed high through the summer. At least three different inhibitors were found in native PAGE. Variation among the trees was high. A transgenic birch carrying the chimeric gene 35SCaMV - proteinase inhibitor II (from potato) did not show higher trypsin inhibitory effect compared to non-transgenic birch.
Promoters. Promoters (35 S, Act 1, win, rol C) connected to the uidA gene were tested on bombarded birch leaves. The variation of GUS-activity within all promoters was high. The wound inducible promoter (win) showed highest activity while the 35S-CaMV promoter had about 20 % of win activity. Gene construct win - uidA was transferred to birch tissue also with Agrobacterium and 30 shoot lines were produced. Wounding of leaves increased GUS-activity systemically 45 fold in greenhouse grown plants.
Early flowering birch. Several early flowering birch clones were micropropagated. These clones will be used to study genetic regulation of birch flowering (Pr Sopanen), and heredity of transferred genes (Pr von Weissenberg). From one of these clones transgenic shoot lines and one embryogenic callus line containing 35SCaMV - uidAINT gene were produced with Agrobacterium mediated gene transfer.
Key words: chitinase, early flowering birch, gene transfer, promoter, proteinase inhibitor.
Correspondence: Ari Pappinen, Department of Plant Biology, FIN-00014 University of Helsinki, Finland
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E-mail: kim.vonweissenberg@helsinki.fi