Improvement Of Hyperparasitic Streptomyces F46 And Streptomyces SC1 By Protoplast Fusion

The excellent characters of the hyperparasitic Streptomyces F46 and the Streptomyces SC1 were tried to integrate into a fusant by protoplast fusion. The study mostly included content as follows:First, the parental strains were identified by two approaches of the classic identify and the molecular taxonomy. Results indicated that the strains F46 and SC1 were classified as Streptomyces griseus and Streptomyces griseoverticillatus.Second, the best situations of protoplast formation and protoplast regeneration of the parental strains were found through experiments. The best conditions of protoplast formation and protoplast regeneration were different in the strains. The parental strains F46 and SC1 were cultured in two-step method. Namely, both are inoculated in the mycelium culture fluid including 20% sucrose respectively, and were cultured for 48 h under the condition 25℃and 150 rpm. Then, they were transferred into the mycelium culture fluid which includes 30% sucrose and 1% or 1.5% glycin under the condition of 20℃, 150 rpm for 48 h. In order to obtain protoplasts of mass and high quality, the strain F46 protoplasts given after combination use of the 10 mg·mL-1 lysozyme and 10 mg·mL-1 snailase at 34℃for 3 h; and the SC1 was treated by 10 mg·mL-1 lysozyme at 34℃for 2.5 h. The protoplasts are not fit to keep at 4℃for long time. The protoplasts of the parental strains F46 and SC1 were inactivated by UV-treatment for 600 s or 360 s, and the both protoplasts were inactivated by heating at 60℃for 45 min or 90 min. Then, in term of single parent or bi-parent inactivate, the protoplast fusion were carried out by PEG 6000 at concentration of 40%, in water bath of 34℃for 10 min. Forty fusants were obtained.Third, Three fusants which had superiority over the parents in many aspects were screened basing on antibiotic activity in vitro. The fusants FSf-11and FSf-13 derived from the fusion between the inactivated protoplast of the strain F46 by heating and the non-treated protoplast of the strain SC1. The fusant FSf-30 was original from the fusion between the protoplast of the strain F46 by heating and the inactivated protoplast of the strain SC1 by UV-treatment. Comparing with the strain SC1, evident inhibiting activity against B. cinerea...