β-1,3-1,4-葡聚糖酶高产菌诱变选育及基因克隆表达

为获得β-1,3-1,4-葡聚糖酶高产菌株,以高地芽孢杆菌YC-9为出发菌株,通过亚硝基胍(nitrosoguanidine,NTG)和低能N+束诱变育种,筛选和选育得到两株突变株N-2-2和10-30s-3,发酵培养60 h,酶活力分别达到28.6 U/m L和36.1 U/m L,分别是出发菌株的2.36倍和2.98倍,与YC-9菌株相比,突变菌株菌体生长下降但发酵产酶量增加。进一步将β-1,3-1,4-葡聚糖酶克隆并在大肠杆菌中成功表达,经过30℃诱导6 h后,胞内酶活力达79.2 U/m L,为出发菌株的6.5倍。

Breeding of High-Yield β-1,3-1,4-Glucanase-Producing Strain by Mutagenesis,and Cloning and Expression of β-1,3-1,4-Glucanase Gene

In order to obtain a high-yield β-1,3-1,4-glucanase producing strain, Bacillus altitudinis YC-9 was used as the
starting strain and mutagenized sequentially with nitrosoguanidine (NTG) and N+ beam. The preliminary and secondary
screening results showed that strain N-2-2 and 10-30s-3 had high ability to produce β-1,3-1,4-glucanase, with a yield of 28.6
and 36.1 U/mL, which revealed a 2.36- and 2.98-fold increase compared with that of the original strain, respectively, while
exhibiting reduced growth rates. Furthermore, the β-1,3-1,4-glucanase gene was successfully cloned and expressed in E. coli.
The intracellular activity was 79.2 U/mL, a 6.5-fold increase compared with the original strain, when induced at 30 ℃ for 6 h.