高密度培养优秀课件.ppt

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1、高密度培养高密度培养第1页，本讲稿共11页Recombinant BoNT/A-Hc as a safe and efficacious vaccine for botulism肉毒神经毒素(BoNT)世界上已知毒性最强的物质之一，共有7种血清型(AG).其中,A型BoNT毒性最强。其重链(HC)的主要作用是与神经细胞膜上相应受体结合，有抗原决定簇，作为疫苗位点。第2页，本讲稿共11页 摇瓶实验 批次培养 菌株的构建Click to ad title in here 123 In this study,the high level expression of the BoNT/A-Hc was

2、 achievedby high cell density cultivation of recombinant Escherichiacoli in a 2-l batch stirred-tank bioreactor.补料分批培养34第3页，本讲稿共11页Bacterial strain and plasmid constructionbont/a-hc genepET28a vectorBL21T7/lac promoterhistidine tagkanamycin resistant gene第4页，本讲稿共11页Glucose 15g/L MgSO4.7H2O 4g/L(NH4)

3、2SO4 3.5g/LK2HPO 4 30g/L KH2PO4 15g/LCitric acid 2g/LNaCl 0.5g/LTrace elements solution 1.5ml/LFeSO4 MnCl2 COSO4 CaCl2 CuCl2 ZnSO4第5页，本讲稿共11页Selection of the best cloneAmong the 10 single E.coli colonies that were grown following transformation with recombinant pET28a,the most suitable recombinant s

4、train was selected using threecriteria:expression level,growth rate and plasmid stability.第6页，本讲稿共11页The plasmid stability of the selected recombinant E.coli strainin grown LB medium during 8 days of subculturing第7页，本讲稿共11页第8页，本讲稿共11页HCDC结果结果第9页，本讲稿共11页Attention Acetate,a well-known growth inhibitor

5、y metabolite for E.coli,is overproduced under anaerobic or oxygen-limiting conditions.DO concentration was maintained above 40%saturation and specific growth rate was fixed at below or 0.2/hduring the fed-batch fermentation.Glucose concentration was also controlled,without any starvation or accumulation of glucose during the process.第10页，本讲稿共11页Thank You!第11页，本讲稿共11页