Callus and cell culture of Tagetes species in relation to production of thiophenes

Abstract

The production of thiophene-biocides by cell cultures invitro was simultaneously investigated with Tageteserecta , T.patula and T.minuta . The calli from which the liquid cultures had to be derived differed between species in the appearance of organoid structure, texture, and colour, Independently of the nutrition of the plants and explants. In particular, the difference between the friability of calli of different species Is obviously related to the expression of the activity of silent genes In only a late phase of the callus andlor the cell suspension culture. Therefore 'origin effects' may eventually determine the suitability of calli to Initiate liquid cultures (Chapter 1). The differences between calli, however, showed that the production of thiophenes In the calli was positively related with the measure of differentiation. Rapidly growing and fine granular cell sus~ pensions, for Instance obtained from smooth calli of T.minuta , did not produce thiophenes (Chapters II and III).Differentiated calli of T.erecta did not provide suitable material to initiate cell cultures In liquid medium. However, minced cauliflower-like calli of T.patula with irregularly occurring small root- or shoot-like differentiations, formed large cell aggregates (3- 8 mm) in liquid media. These cell aggregates accumulated non-polar thiophenes and released spontaneously relatively high amounts of a water-soluble thiophene (BBTOH) in to the medium (Chapter VII). Apparently, the Increased morphological dedifferentation of calli runs parallel with a decreased production of thiophenes In the cell suspensions derived from them. The long-term accumulation of thiophenes In cell aggregates and the release of such compounds into the mediom open perspectives for the commercial production of such compounds under fermentor conditions.Embedding of the fine granular suspension cells of T.minuta in alginate resulted in the release of secondary metabolites into the liquid medium, but did not provide adequate conditions to reinitiate the production of thiophenes (Chapter V). In contrast, naturally formed cell aggregates which can be considered as a natural system of entrapment, as formed by T.patula cells, obviously provide suitable conditions for the production of thiophenes.Genetic transformation of intact Tagetes by means of infection with wild-type and mutant strains of Agrobacteriumtumefaciens and A . rhizogenes , induced neoplastic outgrowth of various organized and unorganized tissues without added growth regulators (Chapter VI). The change in this potential may be related to an altered synthesis of endogenously formed phytohormones. The species-dependent relationship between morphological differentiation and thiophene production persisted In all transformed tissues examined.In conclusion, the results of the present experiments on thiophene production in cell cultures of Tagetes species support the view that, despite the totipotency of plant cells (Chapter IV), major differences exist between closely related species in the ability to serve as a biotechnological unit invitro . Consequently, extensive research to adapt a certain recalcitrant plant species for plant cell biotechnology should be avoided by looking for a better producing species.</TT