耐热α-淀粉酶产生菌枯草芽孢杆菌抗噬菌体的选育

Breeding of Phage-resistant Mutants of Bacillus subtilis Producing Thermostable α-amylase

[目的]选育出耐热α-淀粉酶产生菌枯草芽孢杆菌抗噬菌体菌株。[方法]从耐热α-淀粉酶产生菌枯草芽孢杆菌菌株HA06的异常发酵液中分离到了2种噬菌体,将其命名为KB011、KB012。以枯草芽孢杆菌HA06为出发菌株,采用噬菌体、紫外线和MNNG复合诱变法选育具有抗性的高产突变株,并对突变株的遗传稳定性和发酵的酶活力进行了考察。[结果]获得了3株抗性株,将其命名为KSB04、KSB08、KSB14,它们在试验浓度下对噬菌体KB011、KB012具有明显的抗性,在整个培养过程中表现正常。而敏感菌株HA06的生长则受到明显抑制,并产生大量的噬菌体,培养液中噬菌体的浓度最高可达1012pfu/ml以上。抗性株KSB04和KSB14有较好的遗传稳定性,在7次传代后酶活力仍在3 500 U/ml以上。[结论]获得了遗传稳定且发酵性能与出发菌相似的2株抗性株KSB04和KSB14。

[ Objective] The aim was to breed the phage-resistant mutants with Bacillus subtilis HA06. [ Method] Two kinds phages were separated from the abnormal ferrnentation broth of Bacillus subtilis HA06, which was named as KB011 and KB012 . With Bacillus subtilis HAO6 as an original strain, the work of Breeding of Wage-resistant Mutants was done by the compound mutagenesis method which the two phages,ultraviolet and MNNG as the rnutagens, and also the genetic stability of the mutant strains and their fermentation enzyme activity were investigated . [ Result] 3 high productive mutant strains with resistance were obtaind,named KSB04,KSB08 and KSB14,which had obvious resistance on phage ICB011 and KB012,while the strain HAO6 was inhibited remarkably by the great generated phages which could reach above 10^12 pfu/ml at highest level in the culture medium.Also, the two resistant strains KSB14 and KSB14 had the better genetic stability,and their enzyme activity was still at above 3 500 U/ml after passage of 7 generations . [ Conclusion] The 2 resistant strains KSBO4 and KSB14 with stable heredity were obtained and their fermentation performance was as same as that of the original strain.