Somaclonal Variation Among Tissue Culture Regenerants of New Zealand-Bred Potato (Solanum Tuberosum) Cultivars

Abstract

Explants of shoot cultures of 5 New Zealand-bred potato (Solanum tuberosum) cultivars Ilam Hardy, Iwa, Rua, Tekau and Kaituna and the line 'Lincoln', grown in vitro have been used to determine the ability of axillary buds, complex explants and mesophyll protoplasts to regenerate plantlets. Intercultivar differences in reaction to nodal multiplication and shoot regeneration have been detected, and multiplication rates of nodal cuttings of six to eight-fold per four-weekly subculture 'were achieved. Using a two-step regeneration protocol, explants were induced to form callus in a medium containing an auxin and a cytokinin. The callused explants were transferred to a shoot-induction medium containing the same cytokinin and gibberellic acid A3. It was found that de novo shoots formed more prolifically on leaflet explants than' on stem explants. Gibberellin was required in the shoot-regeneration medium in the presence of benzyladenine, but was less influential when natural cytokinins were used. Plants were regenerated from mesophyll protoplasts derived from in yitro shoot cultures of cvs llam Hardy, Kaituna, and the line Lincoln. A range of variation was detected among regenerated plants when they were outplanted, with alterations in growth and vigour, haulm characteristics and tuber characters. Analyses of phenotypic characters among glasshouse-grown regenerants of Ilam Hardy (To generation) from callus explants revealed approximately 3% with gross aberrations, obvious phenotypic variations or chlorophyll mosaics. This ratio was lower than the near 40% of protoplast-derived Ilam Hardy clones with gross aberrations or that were identified as phenotypic variants. None of the Iwa (To) somaclones were either aberrant or showed chlorophyll mosaicism. No variation was detected in plants grown from axillary bud cultures. A small number of regenerants showed improvements in the first tuber (T1) generation, grown under field conditions. Improvements were observed in the major agromonic characters required by the breeder, i.e. yield and quality, that had not been expressed in the parent plants. The plants bearing these changes had been regenerated from callus using a variety of growth regulators, (auxin, cytokinin and gibberellin) after varying periods on the shoot induction medium. The field trials established that approximately 95% of the Ilam Hardy T1 somaclones and 82% of the Iwa T1 somaclones were genetically stabe, expressing no new characters. Electrophoretic studies were used to identify cultivars and to identify the regenerants with biochemical variation in their tuber sap. Minitubers were formed in vitro from three-week-old shoot cultures by overlaying a liquid tuber-inducing medium containing an amino acid supplement and high levels of benzyladenine and sucrose. Minitubers were ready to harvest eight weeks after application of the induction medium. Sprout, induction commenced over a period of two months. Glasshouse and field growth of minitubers produced plants which were capable of generating yields similar to the parent cultivar, albeit with a higher number of smaller tubers. Plants of the cultivars and regenerants were subjected to zoospores of Phytophthora infestans (late blight) to assess their levels of resistance and whether any regenerants had altered susceptibility to the pathogen, both in vitro and in vivo. No alterations were detected between the parent cultivars and the somaclones. Minitubers and in vitro shoot cultures were used to store pathogenic cultures of Phytophthora infestans, to ensure continuous availability for testing.