摘要
旨为从高寒草地燕麦根际定向筛选解植酸磷微生物资源,筛选促生潜力菌株,分析植酸酶编码基因。采用国际植物研究所磷酸盐生长培养基(NBRIP)分离及筛选菌株,16S rRNA基因鉴定其分类地位,并测定菌株植酸酶活性及促生特性,结合简并PCR和高效热不对称交错PCR(hiTAIL-PCR)扩增植酸酶基因完整序列,并进行生物信息学分析及异源表达。共获得107株菌株,其中51株能在NBRIP培养基上形成清晰溶磷圈,鉴定为2门10科11属,以假单胞菌(Pseudomonas)为优势菌。14株不同种假单胞菌均检测出植酸酶活性,具有溶解有机/无机磷、分泌IAA(3-indoleacetic acid)、固氮及拮抗植物病原菌的促生活性。获得了3株菌株的植酸酶(PHY65、PHY101和PHY131)序列,预测为β-螺旋植酸酶(β-propeller phytases,BPPhy)家族蛋白,其中重组PHY65的比活性为28.2 U/mg。研究结果可为解磷生物菌剂的研发与利用提供优良菌株资源,为植酸酶的生产应用提供理论基础。
This work aims to directly screen the microbial resources dissolving phytate phosphorus in Avena sativa rhizosphere from alpine grassland and plant growth promoting bacteria(PGPB)and to analyze the genes encodingphytase.The strains were isolated and screened using national botanical research institute's phosphate growth medium(NBRIP)and the 16S rRNA gene sequencing were used to identify the strains.The dominant strains were used to determine phytase activity and growth-promoting characteristics.The complete sequence of phytase was amplified by degenerate PCR and high-efficiency thermal asymmetric interleaved PCR(hiTAIL-PCR),and bioinformatics analysis and heterologous expression of new phytase.A total of 107 strains were obtained,of which 51 strains formed a clear phosphate solubilizing circle on NBRIP medium,and the strains belonged to two phyla,10 families and 11 genera,which Pseudomonas(52.94%)were the predominant bacteria.Fourteen Pseudomonas sp.were detected to have phytase activity,and to have the growth-promoting ability of dissolving organic/inorganic phosphorus,secreting 3-indoleacetic acid(IAA),fixing nitrogen and antagonizing phytopathogenic fungi.Three BPPhy(PHY65,PHY101 and PHY131)sequences of strains were cloned from fourteen Pseudomonas sp.,which were predicted to be β-propeller phytases(BPPhy)family proteins,and the specific activity of recombinant PHY65 was 28.2 U/mg.The results may provide excellent strain resources for the development and utilization of phosphate solubilizing agents,and provide a theoretical basis for the production and application of phytase.
作者
李琦
杨晓蕾
李晓林
申友磊
李建宏
姚拓
LI Qi;YANG Xiao-lei;LI Xiao-lin;SHEN You-lei;LI Jian-hong;YAO Tuo(College of Grass Industry,Gansu Agricultural University,Key Laboratory of Grassland Ecosystem Education,China-America Grassland and Animal Husbandry Sustainable Development Research Center,Lanzhou 730070;Jinchuan District Agricultural Technology Popularization Service Center of Jinchang City,Jinchang 737100)
出处
《生物技术通报》
CAS
CSCD
北大核心
2023年第3期243-253,共11页
Biotechnology Bulletin
基金
国家重点研发计划(2019YFC0507703)。
关键词
燕麦
植酸磷
根际促生细菌
16S
rRNA基因鉴定
植酸酶基因
Avena sativa
phytate phosphorus
plant growth promoting bacteria
16S rRNA identification
phytase gene
