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Myxobacteria is a Gram-negative bacteria, it has a complex multicellular behavior, fiber pile sac fungus is one of the sticky bacteria capable of degrading cellulose taxa. Fiber heap capsule bacteria capable of producing a variety of anti-fungal, anti-bacterial and anti-tumor effect of the biologically active substance, wherein the macrolide antibiotics to inhibit the growth of tumor cells by stabilizing microtubule Epothilone particularly noticeable. Epothilone class of drugs so far have been successfully listed, has played a very important role in the fight against tumor cells. However, the production of the Epothilone is mainly dependent on the chemical synthesis method, by contrast, microbial production method has not been widely used, the main limitation is due to its producing bacteria fiber heap capsule bacteria fermentation lower production levels, Breeding yielding fiber heap capsule bacteria to improve the epothilone yield has industrial significance. Fibrous the heap capsule bacteria So 0157-2 efficient Strainfrom, screened epothilone A, B production have greatly improved the mutant LG3111776, its high-yielding strains of epothilone A and B yield about 5 times and 3 times the original strain. Next, we epothilone antitumor mechanism carried out in-depth analysis of epothilone A synergy between B mixed formulations epothilone role of apoptosis, autophagy phenomenon has been described. Text Strainfrom fibrous the heap capsule bacteria So 0157-2 as the original strain, the introduction of the pH extreme environmental stress Breeding starting strain acclimation and enrichment predominant strains on four tolerance strains by high-performance liquid Chromatography its fermentation broth, epothilone A, B, Epothilone standard curve fitting analysis of the changes in its production, screening the predominant strains of LG31 production increased. LG31 again as the original strain, traditional UV mutagenesis and diethyl sulfate mutagenesis, combined with the screening of a 96-well plate high-throughput screening method, high performance liquid chromatography verify further screened mutant LG31-11-7 . Subsequently, the combined method using of UV mutagenesis reasoning breeding strain LG31-11-7, respectively, to tolerate high concentrations of the precursor sodium acetate and tolerate high concentrations of end products two angles efficient breeding and with biological indicator method the rapid screening thin-layer chromatography and TLC positive mutants, the ultimate high-yielding strains LG3111776. Optimization of fermentation conditions on overproducing strain LG3111776 to the steady increase in the production of epothilone eventually stabilized at about 43.1 mg / L and 22.9 mg / L, compared with the initial strain production increased more than 5 times and 3 times more. Not only for protoplast fusion the strain diversity parental good strain resources and industrial production increased. When Epothilone acting on multiple strains of tumor cell lines, Epothilone exhibit good anti-tumor activity. PC3, HeLa, KB, K562, in MCF-7, U87 epothilone most sensitive and epothilone A the B mixed formulations role after found a unique synergies. Through the cell cycle, apoptosis, cell growth and Xcelligence monitoring are three ways to further validate this synergistic effect. We infer the possible reasons of such synergies epothilone A, ion charge attracting force between the B competition between the role of the epothilone tumor cell surface protein to achieve different degrees of saturation. Further, when the epothilone A, B role in the U87/LC3 of tumor cells when fluorescence microscope observation Epothilone can cause LC3 protein (microtubule associated protein) typicality change point poly-shaped, said Ming Aibo neomycin U87 cells can induce autophagy, autophagy is one of the programmed cell death pathways. By the GFP fluorescence transfection tag U87 cells LC3 protein, we can clearly observed autophagy caused by the dot-like structures in the cytoplasm. DAPI-stained nucleus process of change from the other hand, the epothilone certain dependence between U87 cells induced apoptosis and autophagy, which will contribute to the mechanism on the Epothilone drugs inhibit the activity of tumor cells analysis.
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