| Spinosyn, a novel microbial broad-spectrum insecticide, was secondary metabolites from the aerobic fermentation of Saccharopolyspora spinosad. Its favorable mammalian and environmental profile, insect selectivity, unique mode of action and outstanding efficacy lead to high benefit in ecomony and society. In this paper, we are aimed to improve the spinosyn-producing strain by genome shuffling, optimization of fermentation process and scale-up of fermentation processes in 2.5 L fermentor.Firstly, the medium composition for spinsyn production were investigated by 'one-variable-at-a-time' approach (OVAT) and orthogonal design. The optimal medium composition was as follows: Glucose 5.5%, Fish meal 1.5%, Corn powder 2.5%, Soybean powder 0.2%, Starch 0.5%, Soybean oil 0.2%, Sodium Glutamate 0.2%, Calcium carbona 0.5%, pH 7.0, the inoculum amount 10%, medium volume 30ml in 250ml shaking flask. And the addition of 0.3% alchol and 0.3% n-butyl alcohol can improve the production of spinsyn by 52%.In this dissertation, the conditions for protoplast preparation, regeneration and fusion were optimized. According to the biosynthesis pathway and the metabolic regulation of spinosad, strain improvement was performed by protoplast mutatation with UV and LiCl, protoplast fusion and genome shuffling. A high spinosyn producing S. spinosad sp.4.d7 was obtained, and its production reached 547 mg·l-1, which was 202% more than that of the parent strain sp.y.5, and 436% more than that of the original strain sp 102-7. Subculture experiment indicated that the hereditary character of Sp.4.d7 was stable. Spinosad fermentation of S. spinosad sp.4.d7 was scaled up in a 2.5L fermentor in the condition of 350 rpm, DO 60%, 28°C, and its production was 428 mg·l-1.
|
PDF Full Text Request