| Agro-antibiotic 2-16 is an agricultural antibiotic with extensive inhibition effect, which was discovered from the culture filtrate of Streptomyces ahygroscopicus var. huangshanensis. Agro-antibiotic 2-16 exhibits strong effects on many pathogenic fungi of crops, and also some extent of inhibition effects on yeast, some G+ bacteria and G bacteria. When agro-antibiotic 2-16 was used to control Sclerotinia sclerotiorum in the field test, the control effect was more than 65%. Before this study, the production of 2-16 is low, with the titre of only 2000u/ml. In order to improve its production, four mutagenic methods were used in this work. At last, a high-yield strain 515# was obtained. By using Plackett-Burman design and Response Surface Analysis provided by SAS software, the cultivation condition of 515# was optimized. The results were as follows:1 .The initial strain 0# (with titre of 2000u/ml) was treated by means of protoplast regeneration, ultraviolet radiation, NTG mutagenesis and low energy C+ ions implantation respectively. Every time, a strain with higher titre acqucired from one step was selected as the starting strain of the next step. No high-yield mutant was found after protoplast regeneration, strain 301# (with titre of 2317u/ml) was got by ultraviolet radiation, strain 442# (with titre of 4084u/ml) was got by NTG mutagenesis, strain 515# (with titre of 6462u/ml) was got by low energy C+ ions implantation. The production of 515# was increased by 223.10% in comparision with that of 0#. Low energy C+ ions implantation contributed most to the improvement of the production among the four methods, which was 53.29%, and NTG was the second one, which was 39.60 %. In this study, the effects of NTG mutagenesis and low energy C+ ions implantation were better than those of protoplast regeneration and ultraviolet radiation obviously, and were effective methods for the mutation and breeding of the agro-antibiotic 2-16 producing strain.2.By using Plackett-Burman design and Response Surface Analysis provided by SAS software, the cultivation condition of strain 515# was optimized. Compound K and compound F were chosen as the major components from nine variables, a regression model equation was obtained after regression analysis performed on the data from RSA experiment, which is: Y=8914.80-0.2749 x2-298.65 x9-170.53x22-19.75 x2 x9-299.78 x92. The optimum concentrations of compound K and compound F were predicted to be 0.018% and 0.03%, respectively, for the maximum antibiotic production, which is 8989 u/ml. The model was validated experimentally by the maximum production of (8952 94)u/ml. The agro-antibiotic 2-16 production was increased by 38.53% when the strain 515 #was cultivated in the optimum medium instead of the initial one.3.The analysis of the culture filtrates of different strains with High Performance LiquidChromatogram indicted that there were two main components--A and B in agro-antibiotic 2-16, andthe content of B was higher than that of A. The contents of A in the culture filtrate of strain 442# and strain 301# were both higher than that of the initial strain 0#, while the contents of B were both lower.However, when it came to strain 515# and 515# in the optimum medium, the case was the opposite one. The contents of B in the culture filtrates of strain 515# and 515# in the optimum medium were both higher than that of the initial strain 0#, while the contents of A were both lower. The content of B has greater impact on the titre of agro-antibiotic 2-16 than that of A, and was the key factor that dominated the titre of agro-antibiotic 2-16.
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