生防菌Act12铁载体合成酶Ser基因的功能

Functional analysis of Ser gene from biological bacterium Act12

摘要:

【目的】通过敲除生防菌Act12铁载体合成酶Ser基因，研究该铁载体在植物防病促生中的作用。【方法】以自杀型质粒pKC1132作为基本载体，两侧为Ser基因上下游片段作为同源交换臂，两个同源臂之间为卡那霉素抗性基因，构建重组质粒pCT12；借助接合转移技术，将该质粒导入Act12，筛选突变株并进行PCR验证；研究Ser基因缺失突变体和野生型菌株Act12在生长速率、铁载体产量、甜瓜种子促生、抗苹果轮纹病菌(Macrophoma kawatsukai)等方面的差异。【结果】经PCR验证及测序均证实Ser基因缺失突变体构建成功。Ser基因突变后，铁载体合成量明显减少，抑制甜瓜种子的萌发及生长，对苹果轮纹病菌拮抗作用降低。【结论】生防菌Act12 Ser合成酶基因参与控制合成的铁载体在对甜瓜种子促生作用和拮抗苹果轮纹病菌中发挥一定作用，为进一步研究Act12防病促生机制奠定基础。

Abstract:

[Objective] To explore the effects of bio-control and growth-promoting of the siderophore involved with Ser gene in Act12, the siderophore synthase Ser gene knock-out mutant was generated. [Methods] For generating deletion mutant, suicide plasmid pKC1132 was chosen as a basic vector, kanamycin resistance gene as a marker, upstream and downstream of Ser gene as homology arms to construct the recombinant plasmid named pCT12. Then, it was transfered into Act12 by conjugal transfer. Putative deletion mutants were verified by PCR. Whereafter, biological analysis was conducted to assess the role of Ser gene in bio-control and growth promoting. [Results] The Ser deletion mutant ?ser was obtained by PCR and sequencing. Results of biological analysis showed that ?ser synthesizes less siderophores compared to wild strain Act12. Furthermore, the seed germination rate of Cucumis melo L and the antagonism against Macrophoma kawatsukai both significant declined. [Conclusion] These results suggest that Ser gene of Act12 has a certain impact on bio-control and growth promoting, which would establish the foundation for further mechanism study.