Chemical Induction With'EMS'and Screening Mutation Resistant To Grey Mildew Disease In Strawberry (Fragaria Ananassa Duch.)

Mutagenized (EMS treated ) calli from leaves of strawberry (Fragaria ananassa Duch.) 'Sachinoka'were screened for resistance to the culture filtrate of grey mildew pathogen Botrytis cinerea Pers. The purpose of the study was to solve the problem caused by grey mildew disease in strawberry industry by obtaining resistant cultivars and privide a new approach for resistant breeding of strawberry.To obtain a reliable, rapid, and an efficient regeneration system of leaf in vitro, various kinds of factors affecting regeneration including dark treatments, leaf ages, colors of plastic films, silver nitrate and hormones were investigated. The results showed that calli reached their highest regeneration when treated in dark for 6 weeks; regeneration abilities of leaves increased along with their age within 30ds; red films induced the most effecient shoot regeneration, followed by yellow and green ones, but purple films completely inhibited regeneration. Ag~+ enhanced the regeneration abilities , and the regeneration rate reached the peak at 4.0mg/L then decreased with the increase of Ag~+. TDZ concentrations from 2.0 mg/L to 6.0 mg/L did not affect regeneration, but regeneration was depressed exceeding the range. Different kinds of auxin had different effects on regeneration. Auxin of indole promoted regeneration, whereas, 2,4-D had negative effects on regeneration. Different concentrations of NAA showed varied effect in the regeneration..EMS was prepared at concentrations of 0.05%, 0.1%, 0.2%, 0.4% and 0.8%, and calluses were treated with these solutions for 0.5h, 1 .0h or 2.0 h.. The results indicated that browning and mortality of explants rose along with the increases of EMS concentration and treatment time. The activities of PPO, SOD,CAT,POD and contents of MDA, total hydroxybenzene and free proline in explants treated with EMS were measured in order to find causes resulting in and controlling browning during the mutation process.The poisonous effects of culture filtrate of strawberry grey mildew were found both on cell and on plant levels through in vitro observation and inoculation in field, so the culture filtrate was used as crude toxin extract for selection pressure of mutation. To determine the selection pressure, different concentrations of culture filtrate were added to subculture medium and the callus were measured. At last, medium containing 80% of the culture filtrate and 10d culture were selected as selection pressure.Mutant calli with resistance to grey meldew toxin were obtained after twice selections. Some of them produced shoots which would be rooted and then transferred to field to test the resistance.