原生质体融合选育冠菌素高产菌株

Breeding of high-yield coronatine-producing strain by protoplast fusion

研究以抗卡那霉素突变株189(KMr、Glu+)和谷氨酸缺陷营养型突变株A4.6.727(Glu-、KM)s为亲本,进行原生质体融合构建冠菌素高产菌株。确定了制备原生质体的适宜条件为溶菌酶浓度0.05mg/mL和酶解时间100min,此条件下原生质体形成率达79.1%,再生率为12.8%。所得融合子经多次转管培养,检测融合子发酵产冠菌素产量,最终筛选到一株高产冠菌素菌株S1893,其冠菌素相对产量比出发菌株提高184.1%和519.0%。

Using strain 189（KMr, Glu＋） and A4.6.727（GIu-, KM^＋） as parents strains, high coronatine-producing strain was constructed by protoplast fusion technology. The optimal conditions of preparing protoplast were as follows： the lysozyme concentration 0.05mg/mL and hydrolysis time 100min. Under these conditions, the efficiency of protoplast formation was 79.1% and the rate of protoplast regeneration reached 12.8%. The coronatine yields were determined after cultured repeatedly. Finally, a strain S 1893 with high yield of coronatine was successfully screened. The relative yield of coronatine was increased by 184.1% and 519.0% compared to those of the original strains, respectively. The results demonstrated that the protoplast fusion technology is one of the effective methods to obtain high-yield strain.