Proceedings of the XLVII Italian Society of Agricultural Genetics - SIGA Annual Congress

Verona, Italy - 24/27 September, 2003

ISBN 88-900622-4-X

Poster Abstract - 5.61

ECTOPIC EXPRESSION OF “BANYULS” INCREASES LEAF TANNIN LEVELS IN LOTUS CORNICULATUS BUT NOT IN MEDICAGO SATIVA

S.M. HESAMZADEH HEJAZI, F. PAOLOCCI, F. DAMIANI

C.N.R. Istituto Genetica Vegetale sez. di Perugia

condensed tannins, forage legumes, genetic transformation, gene expression

Modification of tannin levels in forage tissue of leguminous plants is essential for increasing palatibility and protein assimilation by ruminants.

From a A.thaliana “trasparent testa” (seed coat tannin free) mutant a structural gene supposed to code for LAR, the enzyme that from the reduction of leucoanthocyanidin produces cathechin, was isolated (Devic et al., 1999) Catechin is the starting element for the condensation of monomeric elements of tannins. Such gene after subsequent investigation (Xié et al., 2003; Abrahams et al., 2002) resulted not coding for a LAR (leucoanthocyanidin reductase) but for ANR (anthocyanidin reductase) giving origin to epicathechin, a catechin enantiomer.

From such observation it was supposed that condensed tannins produced in Arabidopsis were different from those of leaf legumes.

From previous transformation in tobacco it was observed that banyuls redirected anthocyanin synthesis to condensed tannins, and flowers lost the characteristic pink color (Xié et al., 2003).

Banyuls was inserted under constitutive promoter in the weak tannin producer Lotus corniculatus genotype S50 (Carron et al., 1994) and in Medicago sativa Regen Sy27 through Agrobacterium transformation. Several transgenic plants were produced from each species. They were tested for transgene copy number and expression. Qualitative and quantitative analysis of condensed tannins was performed on leaves and flowers. Medicago transgenic resulted identical to the mother plant therefore did not show any tannin on leaves, and flowers reacted with tannin specific reagent producing a pink stain not characteristic of tannins, both in transgenic and controls. Leaves of lotus showed increased levels of tannin respect to the starting plant and were high variable for tannin levels in strict relation with transgene copy number. One transgenic did not accumulated any tannins in leaves and contained several copy of banyuls, possibly a silencing mechanism occurred.

Preliminary results of such work indicated that in forage legumes Banyuls can not induce autonomously tannin accumulation neither in leaves nor in flowers however if the tissue is committed for tannin accumulation banyuls can strongly increase such levels. The theoretical consequences of such observation are that a) epicatechin can substitutes cathechin in tannin formation, b) the low level of tannins can derive from the low levels of monomeric elements c) that an enzymatic complex for condensation and accumulation of tannin should be active, otherwise such elements are not accumulated.

Bibliography

Abrahams et al (2002) Plant Physiol 130: 561-576

Carron et al. (1994) Theor Applied Genet 87:1006-1015

Devic et al. (1999) Plant Journal 19:387-398.

Xié et al. (2003) Science 299:396-399