Streptomycin resistance-aided genome shuffling to improve doramectin productivity of Streptomyces avermitilis NEAU1069.

Genome shuffling is an efficient approach for the rapid engineering of microbial strains with desirable industrial phenotypes. In this study, a strategy of incorporating streptomycin resistance screening into genome shuffling (GS-SR) was applied for rapid improvement of doramectin production by Streptomyces avermitilis NEAU1069. The starting mutant population was generated through treatment of the spores with N-methyl-N'-nitro-N-nitrosoguanidine and ultraviolet (UV) irradiation, respectively, and five mutants with higher productivity of doramectin were selected as starting strains for GS-SR. Finally, a genetically stable strain F4-137 was obtained and characterized to be able to yield 992 ± 4.4 mg/l doramectin in a shake flask, which was 7.3-fold and 11.2-fold higher than that of the starting strain UV-45 and initial strain NEAU1069, respectively. The doramectin yield by F4-137 in a 50-l fermentor reached 930.3 ± 3.8 mg/l. Furthermore, the factors associated with the improved doramectin yield were investigated and the results suggested that mutations in ribosomal protein S12 and the enhanced production of cyclohexanecarboxylic coenzyme A may contribute to the improved performance of the shuffled strains. The random amplified polymorphic DNA analysis showed a genetic diversity among the shuffled strains, which confirmed the occurrence of genome shuffling. In conclusion, our results demonstrated that GS-SR is a powerful method for enhancing the production of secondary metabolites in Streptomyces.