丙酮丁醇梭菌(Clostridium acetobutylicum)原生质体融合育种研究

为了提高丙酮丁醇梭菌(Clostridium acetobutylicum)的丁醇耐受能力和培养基总糖产丁醇的转化率,通过原生质体融合的方法,研究了溶菌酶浓度及其作用时间、再生培养基种类、55℃条件下菌体致死时间、不同PEG分子量以及作用时间、Ca^2+和Mg^2+不同的添加量对丙酮丁醇梭菌原生质体制备、融合、再生的影响,得到了一套比较系统的丙酮丁醇梭菌的原生质体融合条件,同时通过气相色谱检测了融合菌的产溶剂能力并计算总糖转化率。结果显示,最终得到的215I菌株的总糖转化率比原始菌株提高了34.7%,产丁醇能力比原始菌株提高了32.2%,并且发现1株融合菌能产生新物质。原生质体融合方法在丙酸丁醇梭菌育种方面有广泛的应用潜力,通过融合得到的菌株为丁醇生产奠定了基础。

Study on Protoplast Fusion and Breeding of Clostridium acetobutylicum

The study was airmed to improve the butanol tolerance of Clostridium acetobutylicum and increase the conversion rate of total sugar in culture medium to produce butanol by the protoplasmic fusion method. The effect of concentration of lysozyme and its action time, the type of regenerative medium, the time of thermal death at 55 ℃, different PEG molecular weights and the action time, Ca2+, Mg2+ additions with different amounts on Clostridium acetobutanol protoplasm preparation, fusion, regeneration was studied. We obtained a set of comparative system of Clostridium acetobutanol protoplasm fusion conditions. And we detected the solvent production capacity of the fusion bacteria by gas chromatography and calculated the total sugar conversion rate. Results showed that, the total sugar conversion rate of the final obtained strain 215I was 34.7% higher than the original strain, and the butanol production capacity was 32.2% higher than the original strain, and one fusion strain was found to produce new substances. The protoplasmic fusion method is promising in the breeding of Clostridium acetobutylicum, and the strains obtained by fusion provide a basis for further experiments.