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FERMENTATION, CELL CULTURE AND BIOENGINEERINGStreptomycin resistance-aided genome shuffling to improvedoramectin productivity of Streptomyces avermitilis NEAU1069Ji Zhang•Xiangjing Wang•Jinna Diao•Hairong He•Yuejing Zhang•Wensheng XiangReceived: 3 December 2012/Accepted: 25 April 2013/Published online: 9 May 2013? Society for Industrial Microbiology and Biotechnology 2013Abstractthe rapid engineering of microbial strains with desirableindustrial phenotypes. In this study, a strategy of incorpo-rating streptomycin resistance screening into genomeshuffling (GS-SR) was applied for rapid improvement ofdoramectinproductionbyNEAU1069. The starting mutant population was generatedthrough treatment of the spores with N-methyl-N’-nitro-N-nitrosoguanidine and ultraviolet (UV) irradiation, respec-tively, and five mutants with higher productivity of dora-mectin were selected as starting strains for GS-SR. Finally,a genetically stable strain F4-137 was obtained and char-acterized to be able to yield 992 ± 4.4 mg/l doramectin ina shake flask, which was 7.3-fold and 11.2-fold higher thanthat of the starting strain UV-45 and initial strainNEAU1069, respectively. The doramectin yield by F4-137in a 50-l fermentor reached 930.3 ± 3.8 mg/l. Further-more, the factors associated with the improved doramectinyield were investigated and the results suggested thatmutations in ribosomal protein S12 and the enhancedproduction of cyclohexanecarboxylic coenzyme A maycontribute to the improved performance of the shuffledstrains. The random amplified polymorphic DNA analysisshowed a genetic diversity among the shuffled strains,which confirmed the occurrence of genome shuffling. Inconclusion, our results demonstrated that GS-SR is apowerful method for enhancing the production of second-ary metabolites in Streptomyces.Genome shuffling is an efficient approach forStreptomycesavermitilisKeywordsscreening ? Doramectin ? Streptomyces avermitilisNEAU1069Genome shuffling ? Streptomycin resistanceIntroductionDoramectin (Fig. 1), sold commercially as Dectomax, is abroad-spectrum macrocyclic lactone anthelmintic that canbe produced by avermectin-producing strain Streptomycesavermitilis in the presence of cyclohexanecarboxylic acid(CHC) [8, 14, 25, 26, 31]. It belongs to the avermectinclass drug family, but possesses better pharmacokineticcharacteristics and efficacy than avermectins [14]. Dora-mectin has already been approved by the Food and DrugAdministration (FDA) as a veterinary drug for the treat-ment of parasites such as gastrointestinal roundworms,lungworms, eyeworms, grubs, sucking lice, and mangemites in cattle, sheep, swine, and others. The propertiesand applications of doramectin have promoted the com-mercial production of this compound. Unfortunately,industrialization of this valuable antibiotic is limited bythe low production in wild-type strain. Although thebiosynthetic pathway of doramectin has been thoroughlyelucidated, and several methods such as site-specificmutagenesis, semi-synthetic DNA shuffling, and combi-natorial biosynthesis have been employed to improvedoramectin productivity, the doramectin yield is stillunsatisfactory because that secondary metabolism is acomplicated network involved in multiple genes and it isdifficult to improve the doramectin yield by modifyingsingle or several genes [25, 26, 31].Classical strain improvement methods such as muta-tion and random selection have succeeded in generatingstrains with industrial potential, but it is a time-consumingJ. Zhang ? X. Wang ? J. Diao ? H. He ? Y. Zhang ?W. Xiang (&)College of Life Science, Northeast Agricultural University,Harbin 150030, People’s Republic of Chinae-mail: xiangwensheng@neau.edu.cn123J Ind Microbiol Biotechnol (2013) 40:877–889DOI 10.1007/s10295-013-1280-8