Study on preparation and regeneration of protoplasts from CoQ10-yield strains | 辅酶Q10产生菌的原生质体制备与再生条件研究

Chinese. 选用辅酶Q10产生菌深红红螺菌和根癌土壤杆菌为出发菌株，系统研究其原生质体制备和再生条件，为辅酶Q10的工业化生产提供技术支持。从菌龄、溶菌酶质量浓度、酶解时间、酶解温度、高渗溶液的选择等，对深红红螺菌和根癌土壤杆菌2株菌进行了原生质体制备和再生条件的研究。① 根癌土壤杆菌原生质体制备和再生的适宜条件为：菌龄13.5 h，高渗溶液为0.8 mol/L NaCl，溶菌酶质量浓度为0.2 mg/mL，酶解时间75 min，酶解温度35 ℃；②深红红螺菌原生质体制备和再生的适宜条件为：菌龄45 h，高渗溶液为0.5 mol/L蔗糖 + 0.02 mol/L的MgCl2•6H2O，溶菌酶质量浓度为3 mg/L，酶解时间60 min，酶解温度37 ℃。得到了根癌土壤杆菌AT-N10和深红红螺菌Rh1.5005适宜的原生质体制备和再生条件。

English. Using Agrobacterium tumefaciens and Rhodospirillum rubrum as original strains, the research studied the proper protoplast preparation and regeneration conditions for providing CoQ10 scale-up process with technical support. The protoplast preparation and regeneration conditions of Agrobacterium tumefaciens and Rhodospirillum rubrum were studied from strain age, enzyme concentration, enzymatic time, enzymatic temperature and the certain of hypertonic solution. ① The proper conditions of protoplast preparation and regeneration for Agrobacterium tumefaciens: 13.5 h aged, hypertonic solution 0.8 mol/L NaCl, enzymatic time 75 min, enzymatic temperature 35 ℃, enzymatic concentration 0.2 mg/mL; ② The proper conditions of protoplast preparation and regeneration for Rhodospirillum rubrum: 45 h aged, hypertonic solution 0.5 mol/L sucrose and 0.02 mol/L MgCl2•6H2O, enzymatic time 60 min, enzymatic temperature 37 ℃, enzymatic concentration 3 mg/L. The proper conditions were obtained for protoplast preparation and regeneration of Agrobacterium tumefaciens and Rhodospirillum rubrum. This would be the basis of protoplast fusion of these two strains.