产淀粉脱支酶菌株XUEX3的分离及其诱变选育

Isolation and Mutagenesis of Isoamylase Producing Strain XUEX3

本研究以铜山烟草(中烟100)生产田根际土壤为材料,通过以支链淀粉作为碳源的固体鉴别培养基的分离培养获得3株产淀粉脱支酶菌株,将其依次编号为XUEX1、XUEX2和XUEX3,采用DNS法测定了3株菌的产酶活性,经过32 h的连续发酵,XUEX3菌株的酶活性峰值达到了16.3 U/mL,XUEX1和XUEX2的峰值均未超过9 U/mL。选取产酶活性最高的XUEX3作为出发菌株进行紫外线诱变选育。结果表明,试验获得的3个正突变菌株XUEX3-1、XUEX3-2和XUEX3-3的产淀粉脱支酶活性与出发菌株相比均有显著提高,其中XUEX3-3的HC值达到了出发菌株XUEX3的3.2倍;经过32 h的连续发酵,XUEX3-1、XUEX3-2和XUEX3-3产酶活性的峰值分别为39.7 U/mL、43.1 U/mL和35.3 U/mL;与对照相比,突变株XUEX3-2的产酶峰值出现的时间提前了4 h,XUEX3-1和XUEX3-3的则推迟了4 h,但经过连续5代的传代培养,正突变菌株XUEX3-1、XUEX3-2和XUEX3-3的遗传稳定。研究结果丰富了产淀粉脱支酶菌株的遗传资源库,为后期的应用开发提供了更多选项。

In this paper,the rhizosphere soil of Tongshan tobacco(Zhongyan 100)production field was used as material,three strains of starch debranching enzyme producing strains were obtained by solid identification medium with amylopectin as carbon source,which were numbered XUEX1,XUEX2 and XUEX3 in turn.The enzyme producing activities of the three strains were measured by DNS method.After 32 hours of continuous fermentation,the peak enzyme activity of XUEX3 reached 16.3 U/mL,XUEX1 and XUEX2 did not exceed 9 U/mL.The XUEX3 strain with the highest enzymatic activity was selected as the starting strain for breeding by ultraviolet mutagenesis.The results showed that the three positive mutants,XUEX3-1,XUEX3-2 and XUEX3-3,which had the highest of enzyme production,had significantly increased the activity of isoamylase compared with the original strain,and the HC value of XUEX3-3 was 3.2 times of that of the original strain.In the continuous fermentation of 32 h,the peak value of enzyme production activity ofXUEX3-1,XUEX3-2 andXUEX3-3were 39.7U/mL,43.1 U/mL and 35.3 U/mL,respectively.Compared with the control,the peak value of enzyme production of XUEX3-2 was 4 hours earlier,and that of XUEX3-1 and XUEX3-3 was 4 hours later.However,after 5 generations of continuous culture,the genetic stability of positive mutants XUEX3-1,XUEX 3-2 and XU-EX3-3 was stable.The results enriched the genetic resource pool of starch debranching strains and provided more options for later application and development.