As part of our program to develop transgenic peach cultivars with improved disease resistance, we showed that grafting of in vitro cultured `Suncrest' peach [Prunus persica (L.) Batsch] tips `onto decapitated stems of Prunus necrotic ringspot virus (PNRSV) infected `Suncrest' shoot cultures, resulted in consistent transfer of virus across grafts as demonstrated by RNA hybridization analysis, suggesting that such a system could be useful for measuring resistance to PNRSV in peach shoot cultures. We have extended these studies to include grafts of `Springcrest' and `Nemaguard' test tips onto `Suncrest' stocks. RNA hybridization analysis showed that PNRSV persists in shoot cultures for 18 months after initiation from PNRSV-infected `Suncrest' trees and after 16 weeks of treatment of 4°C in the dark, suggesting that a supply of infected shoot cultures could be maintained for repeated use. Graft success rates for grafts of `Springcrest' onto `Suncrest' and `Nemaguard' onto `Suncrest', equaled or exceeded success rates for `Suncrest' onto `Suncrest'. Virus was transmitted from infected stocks into `Suncrest', `Springcrest', and `Nemaguard' test tips by 2 weeks in most successful micrografts. There was no significant difference in the virus concentrations among the three scions at 2, 4, and 6 weeks after grafting, suggesting that there is equal efficacy of virus transfer through grafts from `Suncrest' to the three cultivars, and that no differences in resistance to PNRSV exist among these cultivars.
Kathleen Heuss, Qingzhong Liu, Rosemarie Hammond, and Freddi Hammerschlag
Kathleen Heuss-LaRosa, Rosemarie Hammond, James M. Crosslin, Christine Hazel', and Freddi A. Hammerschlag
In vitro micrografting was tested as a technique for inoculating peach [Prunus persica (L.) Batsch] shoot cultures with Prunus necrotic ringspot virus (PNRSV). Cultured `Suncrest' shoots derived from a naturally infected tree (as indicated by ELISA testing) maintained virus in vitro, with virus concentrations in growing tips and folded leaves being several times those of fully expanded leaves. Infected shoots served as graft bases and source of the virus. Grafted tips were derived from `Suncrest' trees that had tested negative for the virus. Leaf samples were collected from the tips following grafting and analyzed for the presence of virus by slot-blot hybridization with a (DIG)-labeled cRNA probe derived from PNRSV RNA 3. Rates of successful grafting ranged from 55% to 73% in three trials and PNRSV was found in all tips analyzed. Virus concentrations approximated those found in source shoots, suggesting that in vitro micrografting should be useful for screening transformed peach shoots for coat protein-mediated resistance to PNRSV. Chemical name used: digoxigenin (DIG).