The Roots of Rye (Secale cereale L.) Are Capable of Synthesizing Benzoxazinoids.

According to current opinion, the first step of benzoxazinoids (BXs) synthesis, that is, the conversion of indole-3-glycerol phosphate to indole, occurs exclusively in the photosynthesising parts of plants. However, the results of our previous work and some other studies suggest that this process may also occur in the roots. In this study, we provide evidence that the first step of BXs synthesis does indeed occur in the roots of rye seedlings. We detected ScBx1 transcripts, BX1 enzyme, and six BXs (2-hydroxy-1,4-benzoxazin-3-one, 2,4-dihydroxy-1,4-benzoxazin-3-one, (2R)-2-O-ß-d-glucopyranosyl-4-hydroxy-(2H)-1,4-benzoxazin-3(4H)-one glucoside, 2,4-dihydroxy- 7-methoxy-1,4-benzoxazin-3-one, 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one glucoside, and 6-methoxy-2-benzoxazolinone) in the roots developed from seeds deprived of the coleoptile at 2 days after sowing (i.e., roots without contact with aerial parts). In roots regenerated in vitro, both ScBx1 transcripts and BX1 enzyme were detected at a low but still measurable levels. Thus, BXs are able to be synthesised in both the roots and above-ground parts of rye plants.

Cucumber (Cucumis sativus L.).

We describe two novel Agrobacterium tumefaciens-based methods of cucumber transformation. The first involves direct regeneration from leaf microexplants selected on kanamycin-containing medium. The second involves regeneration from a long-term established embryogenic suspension culture emitting green autofluorescence (GAF) and selection on medium containing hygromycin. In the latter method, GAF was used as a reporter, thereby allowing a simple and reliable identification of transgenic cells with a high regeneration capacity. (No false positives were observed.) The transformation efficiency in the leaf microexplants fluctuated from 0.8 to 6.5% of the primary explants, whereas in the embryogenic suspension-cultured cells it varied from 6.4 to 17.9% of the aggregates. In the GAF method, the step involving the elimination of the Agrobacterium cells by antibiotics could be omitted; however, this reduced the transformation efficiency to about 3%. The time required from inoculation to regenerated plant in the greenhouse was the same for both methods, but the GAF method required more preinoculation time than the leaf microexplant method.

Structural and ultrastructural analysis of Solanum lycopersicoides protoplasts during diploid plant regeneration.

Light, fluorescence and electron microscopy were used to analyse the structural properties of protoplasts obtained from established suspension culture of Solanum lycopersicoides Dun, composed of meristematic cell aggregates. Four types of protoplasts were distinguished immediately after isolation: (1) mononuclear; (2) polynuclear, (3) anuclear and (4) homogeneous protoplasts. Only mononuclear protoplasts were capable of complete cell wall regeneration and mitotic division. Other types of protoplasts were eliminated during culture. Three phases were distinguished in the developing protoplast culture: (1) the elimination phase during which protoplasts damaged during isolation underwent complete degradation; (2) a phase of intense division during which both mitotic cell division and amitotic nuclear division took place; and (3) a stabilization phase leading to the formation of suspension culture. The cell suspension culture obtained from protoplasts was capable of regenerating diploid plants.