萎缩芽胞杆菌CKL1促盐胁迫下燕麦生长活性及其功能基因分析

Avena sativa growth-promoting activity of Bacillus atrophaeus CKL1 under salt stress and the functional genes

摘要:

【背景】青海省特殊生境孕育了特殊微生物资源。【目的】探究适合生活于高原生境的芽胞杆菌菌源。【方法】采用平板对峙法、显色法对萎缩芽胞杆菌(Bacillus atrophaeus) CKL1的拮抗、产吲哚乙酸活性进行测定，并检测耐低温、耐盐性及菌株对盐胁迫下燕麦品种(Avena sativa) “青燕1号”种子萌发、幼苗生长效应及叶绿素、脯氨酸、丙二醛的含量变化，利用二代测序技术对菌株进行基因组测序并分析相关功能基因。【结果】菌株CKL1对禾谷镰孢菌(Fusarium graminearum)、锐顶镰孢菌(Fusarium acuminatum)表现出显著的拮抗活性(抑菌圈直径>15 mm)；与Salkowski比色液反应变红，能在NaCl浓度为13%的LB培养基及4 ℃低温下生长，表现出一定的产吲哚乙酸、耐盐及耐低温活性；盐胁迫下，菌株CKL1对“青燕1号”种子萌发及幼苗生长具有显著促进作用，叶绿素及脯氨酸含量显著增加，丙二醛含量下降，增强了燕麦的抗盐性。菌株CKL1基因组全长为14 281 280 bp，与GO功能数据库比对注释到3 303个功能基因；基因组编码与脂肽类化合物iturin、surfactin合成及与吲哚乙酸合成相关基因，编码与逆境应答相关Na⁺/H⁺逆向转运蛋白及脯氨酸、甜菜碱等渗透调节物质合成相关基因簇，编码参与高盐及低温胁迫应答转录调控因子的关键基因。【结论】本研究为利用芽胞杆菌促盐胁迫下燕麦生长、提高燕麦抗盐性提供了优异菌株及理论依据。

Abstract:

[Background] The unique eco-environment in Qinghai province allows the growth of special microbial resources. [Objective] To explore Bacillus resources that can tolerate the plateau environment. [Methods] Antagonistic activity and indole-3-acetic acid (IAA) production of Bacillus atrophaeus CKL1 were determined by plate confrontation and chromogenic method, respectively. Moreover, the low-temperature tolerance and salt resistance of CKL1, as well as the influence of CKL1 on seed germination, seedling growth and the content of chlorophyll, proline, and malondialdehyde of Avena sativa 'Qingyan 1' under salt stress were detected. Then, the whole genome of CKL1 was analyzed by next-generation sequencing and the functional genes were dissected. [Results] CKL1 significantly antagonized Fusarium graminearum and F. acuminatum (inhibition zone diameter >15 mm). The reaction solution of CKL1 and Salkowski reagent turned red and the strain grew in LB medium with 13% NaCl and at 4 ℃, indicating that it could produce IAA and was tolerant to salt and low temperature. CKL1 significantly promoted seed germination and seedling growth of 'Qingyan 1' under salt stress, significantly increased the content of chlorophyll and proline, decreased the content of malondialdehyde, and enhanced the salt resistance of 'Qingyan 1'. The genome of CKL1 was 14 281 280 bp, and 3 303 functional genes were annotated against GO. The genome encodes genes related to the synthesis of lipopeptides iturin and surfactin, and the synthesis of IAA, gene clusters related to the synthesis of osmoregulation substances such as proline and betaine, and the Na⁺/H⁺ antiporter in stress response, as well as the key genes encoding transcriptional regulators involved in response to high salt and low temperature. [Conclusion] The study lays a theoretical basis for using Bacillus to promote the growth of A. sativa under salt stress.