Study On The Protoplast Fusion Of Microherbicide Strains

The weed chemical control was the symbol of the modern agriculture development.It was an important measure to save the cost and improve the efficiency of the agriculture, as well as favor the mechanized operation. However, the pesticide pollution was the most serious organic pollution involved the largest areas, the application of pesticide was 500-600 thousand ton and covered 280 million hm2, among which the use of herbicide was 125-150 thousand ton per year. 80% pesticide was introduced into the environment directly on average, diffused into the air and the soil, as well as polluted water both on surficial and under ground. No matter the agricultural products for civil market or export, the standard for the pesticide residue is more strictly, the large margin of reducing use of pesticide was the trend of agriculture development in future. Expanding micoherbicide, biologically control the weeds would reduce large range use of the pesticides which contributed to the safty of the ecology and the food, even the sustainable control of deleterious biology.Exserohilum monoceras (EM) and Curvularia lunata (CL) were isolated by the China National Rice Research Institute which were fungal pathogens of the barnyardgrass from the natural infected the weed plants with the potential mycoherbicide's strains. EM strain had high virulent to barnyardgrass but low spore yield, while the CL strain had high spore-yield but low virulent to the weed. The objectives of this experiment were to breed a new strain through protoplast fusion between the EM and CL which could provide high spore yield and richer toxins, as well as to the favor of industry production.The major results of the experiment showed as follows:1. The strains of EM and CL with antibiotic resistance were obtained by UV-treatment on conidia of EM and CL and screen in Nystatin and Cycloheximide media.2.The optimum conditions were determined to preparation protoplast, which included the hypha of EM and CL obtained after stilly cultivation for 24 h and 18 h respectively, the protoplasts given after combination use of the lytic enzymes of 2% snailase and 2% Lywallzyme at 30℃ for 4 h, the regeneration of protoplasts screened the Re3 media. Protoplast fusion were successful that the protoplast of CL inactivated by UV-treatment for 4 min, and the protoplast of EM inactivated by heating at 55℃ for 5 min, as well as enhancing the fusion process by PEG of 40% at pH 7.5. Three fusants of FK F2 and Rc were obtained.3.The morphology of the fusants colony and the growth velocity of the fusants were significantly different from the parent strains. The DNA polymorphism of the fusants was studied with RAPD-PCR. The characteristic of parent DNA bands was identified from the fusant Rc strain. The barnyardgrass control ability of the fusant F2 was significantly higher than that of CL strains, and similar to that of EM.