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采用抑菌圈法筛选出12株对绿色木霉有明显抑制作用的生防细菌,其中抑菌效果较好的为12C2-4和MS82菌株。通过对峙培养法发现MS82菌株对双孢蘑菇菌丝生长几乎没有影响。通过API20 NE、API50 CH生理生化指标测定及基于16S r DNA基因片段的系统进化分析对MS82菌株进行分类鉴定,结果显示MS82属于假单胞菌(Pseudomonas sp.)。为了明确MS82中参与抗真菌活性相关的基因,利用Tn5转座子随机插入的方式构建MS82菌株的突变体库,获得完全失去绿色木霉抗性的突变体MS82MT31。通过对抗菌失活突变体中突变基因定位,发现突变基因为rpo B基因,与已知菌株Pf0-1的rpo B基因相似性为96%,说明rpo B基因与MS82抑菌活性物质合成密切相关。
Twelve strains of biocontrol bacteria with obvious inhibition on Trichoderma viride were screened by the inhibition zone method. Among them, 12C2-4 and MS82 were the best antibacterial agents. By confrontation culture method found MS82 strains of Agaricus bisporus mycelium growth almost no effect. The MS82 strain was classified and identified by API20 NE, API50 CH physiological and biochemical indexes and phylogenetic analysis based on 16S r DNA gene fragment. The results showed that MS82 belong to Pseudomonas sp. In order to identify the genes involved in antifungal activity in MS82, a mutant library of MS82 strain was constructed by random insertion of Tn5 transposon to obtain mutant MS82MT31 which completely lost the resistance to Trichoderma viride. The results showed that the mutant gene was rpo B gene, and the similarity with the rpo B gene of the known strain Pf0-1 was 96%, which indicated that rpo B gene was closely related to the synthesis of antibacterial substance MS82 .