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Infection of Protoplasts Derived from Potato Shoot Cultures with Potato Virus X

Department of Pure and Applied Biology, Imperial College of Science and Technology, London SW7 2BB
and¹ Ministry of Agriculture, Fisheries and Food, Harpenden Laboratory, Hatching Green, Harpenden, Herts. AL5 2BD, U.K.

Because it contains the Nx gene, potato cv. ‘Maris Piper’ reacts hypersensitively to inoculation with group three strains of potato virus X (PVX). However, protoplasts prepared from aseptically grown shoot cultures of potato cv. ‘Maris Piper’ were infected reproducibly with either PVX or PVX RNA using inocula containing polyethylene glycol. The extent of infection of protoplasts was assessed by fluorescent antibody staining, and by assaying extracts for antigen by ELISA, for infectivity and for PVX RNA content by cDNA hybridization. Infection occurred in 43 to 65% of the protoplasts infected with PVX and in 57 to 70% of the protoplasts infected with PVX RNA. In comparison, when high quality protoplasts were obtained from leaves of pot-grown plants 55 to 65% became infected when inoculated with PVX RNA. However, protoplasts obtained from leaves of whole plants varied greatly in both quantity and quality, whereas shoot cultures reproducibly gave high quality protoplast preparations in large numbers. In protoplasts from either source, yields of progeny virus were between 10 and 30 pg per infected protoplast. The multiplication of a group 3 strain of PVX in protoplasts containing the Nx gene did not induce necrosis and confirms that hypersensitivity is not always expressed in isolated protoplasts.

Keywords: potato virus X, shoot cultures, protoplasts, Nx gene

Received 19 December 1984; accepted 8 February 1985.