Chemical and genetic characterization of calli derived from somatic hybridization between tansy (Tanacetum vulgare L.) and pyrethrum (Tanacetum cinerariifolium (Trevir.) Schultz-Bip.)

Pyrethrum (Tanacetum cinerarifolium (Trevir.) Schultz-Bip.) produces enviro nmentally benign pesticides, the pyrethrins, and tansy (Tanacetum vulgare L .) lower terpenes of variable biological effectiveness. As an approach to i mprove the oil content and composition of tansy for enhanced biological act ivity, a somatic hybridization technique between tansy and pyrethrum was es tablished. About 1 x 10(6) of leaf-mesophyll protoplasts of both species we re mixed and fused with a solution containing 15% polyethylene glycol. Ligh t-green and yellowish calli developed from the fusion experiments. The fusi on-derived calli grew vigorously on MS medium supplemented with 6.4 mgl(-1) of BAP, 0.8 mg l(-1) of NAA, and 30-40 g l(-1) of glucose. Nuclear DNA con tent, RAPD patterns, and volatile compounds were analyzed to determine the hybridity of the calli. The nuclear DNA content of the tansy and pyrethrum genotypes, and the protoplast-derived calli of tansy were 6.41, 7.39, 13.84 , and 8.11 pg, respectively. The nuclear DNA content of individual calli de rived from the protoplast fusion between tansy + tansy ranged from 8.84 (F4 3A) to 19.59 pg (F43C) while those of the tansy + pyrethrum fusions were 10 .66 (F46A) and 31.87 pg (F46B). Using four 10-mer primers a total of 56 RAP D-PCR fragments were produced. The distance matrices of fragments were calc ulated by average linkage cluster analysis. Two visually separated clusters were observed. One cluster consisted of the two tansy genotypes and the fu sion-derived callus F43A; the other consisted of pyrethrum and fusion-deriv ed calli F46B and F46C. Volatile compounds, such as decadienal, artedouglas ia oxide, heptadecane, syringaldehyde and coniferyl alcohol, analyzed by ga s chromatography mass spectrometry, were found only in the protoplast fusio n-derived calli F43A and F46B. Several less volatile compounds were also de tected only in fusion calli. Hexadecanoic and linoleic acids were common to fusion-derived calli and tansy, and one unknown compound to fusion-derived calli and pyrethrum. Pyrethrins I and II were detected from pyrethrum, but not from the fusion-derived calli. The additive nuclear DNA content of pro toplast fusion-derived calli and the results of the RAPDs suggest that inte rspecific fusions had occurred. The small number of volatile compounds dete cted from both the fusion calli and from the donor species indicates that t he unorganized callus tissue is unable to produce tissue-specific volatile compounds.