Optimisation of Turnip (Brassica Rapa) Protoplast Culture for Cauliflower Mosaic Virus Transformation

Abstract

Cauliflower Mosaic Virus is one of the rare plant DNA viruses and consequently a potential vector for genetic engineering (1). Natural infection and proliferation, as judged from ‘mosaic symptoms’, is restricted to a relatively narrow host range, including predominantly members of the Crucifereae. One of the most effective hosts for CaMV is turnip (Brassica rapa ssp. sativa var.rapifera). Natural virus transmission is via aphids and experimental infection via mechanical wounding of leaf epidermis. CaMV spreads systemically through the plant starting from the infection site. (2). We are interested in developing, as an alternative to the Agrobacterium tumefaciens mediated transformation system, a CaMV-based transformation system which makes use of the strong CaMV promotor capacity and the high copy number per cell. One of the basic experimental prerequisites for this aim is a routine protocol for efficient development of proliferating, protoplast-derived cell cultures. The development of such a protocol is the aim of the present study.