水稻根系内生固氮菌的分离鉴定及其促生作用研究  

Isolation,identification and growth promotion of endophytic nitrogen fixing bacteria from rice roots

作　　者：董萌 施龙清[1] 解振兴[1] 连玲[1] 吴春珠[1] 张居念[1] 张数标[1] 姜照伟[1] DONG Meng;SHI Longqing;XIE Zhenxing;LIAN Ling;WU Chunzhu;ZHANG Junian;ZHANG Shubiao;JIANG Zhaowei(Rice Research Institute,Fujian Academy of Agriculture Sciences,Fuzhou,Fujian 350018,China)

机构地区：[1]福建省农业科学院水稻研究所,福建福州350018

出　　处：《西北农林科技大学学报（自然科学版）》2023年第1期31-39,共9页Journal of Northwest A&F University(Natural Science Edition)

基　　金：福建省自然科学基金项目(2020J011353);福建省农业科学院自由探索科技创新项目(ZYTS2019001);福建省农业科学院科技创新团队项目(CXTD2021002-2)。

摘　　要：【目的】研究不同生育期水稻根系内生固氮菌的多样性,分离鉴定内生固氮菌并筛选具有促生作用的菌株,为水稻生产中固氮菌的应用提供理论依据。【方法】采用无氮培养基培养方法,从拔节期和抽穗期佳辐占根系中分离内生固氮菌,通过扩增16S rRNA方法进行分子鉴定和系统进化分析;采用透明圈法进行溶磷解钾能力验证;采用乙炔还原法测定固氮酶活性。将分离的内生固氮菌株单独接种盆栽水稻幼苗,测定其对株高、根长、鲜质量的影响;将内生固氮菌混合菌液接种水稻品种佳辐占和台粳8号,于齐穂期测定倒1、倒2和倒3叶片的叶绿素含量(SPAD值)。【结果】从拔节期和抽穗期水稻根系中分别分离出10和20株内生固氮菌,且两个时期水稻根系内生固氮菌群多样性存在差异,拔节期以克雷伯氏菌属为主,而抽穗期以假单胞菌属为主。筛选出7株具有溶解无机磷能力的菌株,分别是J3、J10、J23、H27、H30、H33和H37。乙炔还原法测定结果表明,分离的内生固氮菌均具有弱固氮活性,其中H33和H40-2菌株固氮能力相对较强。将固氮菌回接水稻幼苗,H40-2和J3菌株对水稻株高、地上鲜质量、根长均有显著影响,可促进水稻生长。将固氮菌混合回接水稻根系,发现佳辐占叶片叶绿素含量与对照差异不显著,而台粳8号显著提高。【结论】分离的内生固氮菌株H40-2和J3在水稻苗期具有明显促生作用,可作为功能性内生固氮菌开发利用。【Objective】This study investigated the diversity of endophytic nitrogen fixing bacteria in rice roots at different growth stages and the strain with growth promoting ability was isolated,identified and screened to provide basis for the application of nitrogen fixing bacteria in rice production.【Method】The nitrogen-free medium was used to isolate the endophytic nitrogen-fixing bacteria,and the 16S rRNA gene was amplified by PCR for molecular identification and phylogenetic analysis.The ability of dissolving phosphorus and potassium was verified by transparent circle method,and the activity of nitrogen fixing enzyme was determined by acetylene reduction method.Endophytic nitrogen fixing strains were inoculated into rice solely at seedling stage,and physiological indexes including plant height,root length and fresh weight were measured.The recombinant endophytic bacteria were inoculated to rice,and the chlorophyll contents(SPAD value)of the top three leaves were measured at heading stage.【Result】A total of 10 strains and 20 strains of endophytic nitrogen fixing bacteria were isolated at jointing stage and heading stage,respectively,and the diversity of endophytic nitrogen fixing bacteria in rice roots was different between the two stages.Klebsiella was dominant in jointing stage and Pseudomonas was dominant in heading stage.Among them,7 strains with the function of dissolving inorganic phosphorus were J3,J10,J23,H27,H30,H33 and H37.The nitrogenase activity of isolated endophytic nitrogen-fixing bacteria was detected,and H33 and H40-2 had strong nitrogen fixation ability.After being inoculated to rice roots,H40-2 and J3 had significant impacts on plant height and fresh weight of rice.The SPAD value of TG 8 increased significantly after inoculating nitrogen fixing bacteria and there was no significant difference in JFZ.【Conclusion】The endophytic nitrogen fixing strains H40-2 and J3 isolated from rice roots significantly promoted the growth of rice at the seedling stage,and can be developed and utilized

关 键 词：内生固氮菌 水稻 固氮酶活性 微生物多样性 

分 类 号：S182[农业科学—农业基础科学] Q939.96[生物学—微生物学]