梭梭根际溶磷菌溶磷关键基因挖掘及其响应磷匮缺的分子机制

Insoluble inorganic phosphate solubilizing is one of the important pathway of phosphate-solubilizing bacteria (PSB) to dissolve phosphorus in soil and promote plant growth. Haloxylon ammodendron, dominantly distributed in desert areas of northwestern China, has strong resistance to drought and infertility. Soluble phosphate (P) is chronically absent in the rhizosphere soil of H. ammodendron. Thus PSB plays an important role for H. ammodendron to adapt to extreme environment. There have been not relevant report on the P solubilization mechanism of rhizosphere PSB colonization in this natural extreme habitats. We have obtained a novel strain M30-35 with the capability of P solubilization from Haloxylon ammodendron rhizosphere, which belonged to the genus Pseudomonas. In this project, we intend to construct mutant library of the strain M30-35 and select inorganic P solubilization deficient mutants using transposon insertional mutagenesis and inverse PCR; then analyze P-solubilizing ability of deficient mutants and its complement strains and evaluate the function of the corresponding inorganic P solubilization genes. Specific functional genes involved in the insoluble inorganic P solubilizing and pathway of the strain M30-35 in response to P depletion will be analyzed by comparing the whole genome between the strain M30-35 and reference strains of the genus Pseudomonas and combining above functional genes. The results will provide the theoretical basis for P solubilization mechanism of PSB in the desert environment, explore a new pathway for efficient use of phosphorus in Plant-PGPR-Soil system and develop a new efficient compound microbial fertilizer to improve crop resistance.

溶解难溶性无机态含磷物质是溶磷细菌（PSB）发挥溶磷作用、促进植物生长的重要途径之一。梭梭是我国西北荒漠特有优势植物，具超强的抗旱耐贫瘠能力，根际土壤可溶性磷长期匮缺，溶磷菌在其适应逆境中发挥着重要作用，目前为止，对这类天然极端生境的根际PSB溶磷作用的相关机制未见报道。申请人从梭梭根际分离的假单胞菌属新种菌株M30-35具有较强的溶磷能力。本项目拟采用转座子插入诱变、反向PCR等技术，构建M30-35突变体文库，筛选溶无机磷缺陷突变体；分析突变体及其互补菌株溶磷特性，明确相应基因的功能；比较M30-35与假单胞菌属PSB参考菌株的全基因组，结合上述具体功能基因，解析M30-35响应可溶性磷匮缺的特异功能基因及其作用途径。研究结果将有助于阐释根际PSB适应特殊生境的溶磷机制，为植物-有益菌-土壤系统的磷高效利用提供新途径，为开发提高作物抗逆性的高效新型复合微生物肥料提供工程菌株。

溶难溶性无机磷是溶磷细菌（PSB）发挥溶磷作用、促进植物生长的重要途径之一。本项目在前期荒漠植物梭梭根际细菌多样性研究的基础上，首先使用优化后的分离纯化法再次从阿拉善右旗巴丹吉林沙漠梭梭根际获得溶磷性能更优的菌株AWH-NS6，并从不缺磷的祁连山云杉林下土壤中获得溶磷性能较好的菌株LT-4，扩充和优化了研究使用的菌株；比较分析了新获得的PSB菌株的生理生化特性及其促生能力，发现新获得的菌株AWH-NS6和菌株LT-4溶磷性能优于原有菌株M30-35；完成了用于后续研究的PSB菌株的全基因组测序分析和相关溶磷功能基因的初步挖掘；基于研究的新发现，采用原核转录组方法，研究了菌株AWH-NS6其溶解难溶性无机磷能力受外源可溶性磷诱导增强的差异表达基因，获得了关键的溶磷响应差异基因；构建了上述菌株的随机突变体库以及菌株AWH-NS6溶磷关键基因pqqE和gcd缺失突变体；综上所述，以上实验结果互为补充，分析全部完成后将填补根际PSB适应特殊生境的溶磷机制相关理论，同时为后续的研究提供了突变菌株库，研究结果也将为植物-有益菌-土壤系统的磷高效利用提供新途径。此外，本研究中获得了大量菌株资源，其中含有若干新种候选菌株，目前已完成其中三株菌株的新种分类鉴定，并发表了相关学术论文，这部分资源将为开发提高作物抗逆性的高效新型复合微生物肥料提供菌株资源。