[ Objective ] The purpose was to research the protoplast production condition and mutation breeding of producing strains of acid pullulanase. [ Method ] The protoplast production and regeneration condition of Halococcus sp. Z1 was researched and its protoplast was used for breeding producing strains of high active pullulanase through ultraviolet mutagenesis. [ Result ] After the strains were cultured for 14 h, it was treated by enzymolysis with 2.8 mg/ml lysozyme at 36 C for 1.5 h, and then the formation rate and regeneration rate of their protoplasts were 87.4％ and 19.4％ resp. Nine mutants were screened out through protoplast mutagenesis by ultraviolet irridiation and the activity of their produced pullulanase was higher than that of origin strain. Among the mutants, the activity of pullulanase produced from D9-08 reached 6.37 U/ml, and compared to that of orgin strain, it was enhanced by 0.96 times. The heritage of D9-08 became stable after 10 passages. [ Conclusion ] It was feasible to breed the producing mutants of acid pullulanase through protoplast mutagenesis by ultraviolet irridiation and the breeding effect of this approach was better.%[目的]对酸性普鲁兰酶产生菌的原生质体制备条件及诱变育种进行研究.[方法]时盐球菌(Halococcus sp.)Z1的原生质体制备和再生条件进行了研究,并对其原生质体进行紫外诱变选育普鲁兰酶高产菌株.[结果]菌体经培养14 h后,在36℃、2.8 mg/ml溶菌酶作用下酶解1.5 h,其原生质体形成率和再生率分别为87.4%和19.4%.采用紫外线对原生质体进行诱变,筛选得到9株普鲁兰酶活性比出发菌株高的突变株,其中D9-08的酶活达6.37 u/ml,比诱变前提高了0.96倍,经10次传代遗传性稳定.[结论]利用原生质体紫外线诱变手段进行酸性普鲁兰酶产生菌的育种是一条可行并具有较好效果的途径.
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