Solubilizing Properties Of The Inorganic Phosphate- Solubilizing Bacteria Isolated From Soybean Rhizosphere Soil

Phosphate-solubilizing bacteria(PSB), a group that can improve soil fertility and soil health by converting insoluble P forms to soluble forms accessible by plants, plays an important role in agriculture P circulate and benefit plant growth development. In this thesis, in order to clear and definite the species of phosphate-solubilizing bacteria strains from the black soil of soybean rhizosphere, the characteristics of dissolving inorganic phosphorus of different PSB. The results are as follows:Strain bacterium was isolated from rhizosphere soil of soybean. The maximum amount of phosphorus solubilization was 558μg/mL, 478μg/mL, 596μg/mL and 586μg/mL, respectively. The Vitek 2 automatic analysis system and 16 S rDNA sequences technology were used to identify the bacterium Pseudomonas sp., Enterobacter sp., Ochrobactrum sp., Klebsiella sp. were named wj1, wj3, wj5 and wj6.On the basis of NBRIP medium, the orthogonal design L25(53)was employed in decision medium optimization from the carbon source, nitrogen source and pH value for strains phosphorus solubilization. The results indicated that the highest capacity to solubilizing phosphate was appeared when strain was supplied with glucose and peptone as carbon source and nitrogen source and the medium pH value was 8. The strain wj6 exhibited higher solubilizing abilities in Ca3(PO4)2, FePO4, AlPO4 as sole source of phosphorus, and maximum amount of phosphorus solubilization were 409.28μg/mL, 60.59μg/mL, 32.90μg/mL, respectively. Furthermore, the solubilizing phosphorus ability of strain wj6 was strongly influenced by environmental pH when FePO4 and AlPO4 were as the phosphorus source. The strain wj6 could dissolve FePO4 and AlPO4 when it was inoculate to pH 8 soluble phosphorus medium. However, no phosphorus solubilizing was found in the pH 3 medium.Organic acids play a major role in release of available P from insoluble phosphorus. The organic acids were identified by GC-MS, In the culture media of 4 strains organic acids 2, 2-dimethoxy propionic acid, α-ketoglutaric acid, 2, 4-hexadienedioic acid, 3-hydroxydecanoic Acid, malic acid, hepta-2,4-dienoic acid, acetic acid, succinic Acid, citric acid were detected. The results of enzyme activity detected. In the process of solubilizing phosphate enzyme activity significantly enhanced.The GDH genes was cloned from wj1, wj3, wj5 and wj6 PSB. Gene segment length is 2007 bp, 2066 bp, 1727 bp and 2391 bp, respectively. Based on the BLAST results, wj1, wj3, wj5 and wj6 were showed 92.30% identity with Pseudomonas brassicacearum subsp. brassicacearum NFM421, 96.51% identity with Enterobacter asburiae L1, 85.12% identity with Ochrobactrum anthropi ATCC 49188, and 99.29% identity with Klebsiella pneumoniae subsp. pneumoniae KP5-1. Comprehensive 12 h, 48 h the changes of GDH genes expression of wj1, wj3 in different phosphorus and pH value culture medium, the results showed that strains in the process of dissolving phosphorus GDH genes were expressed, the same bacteria in different phosphorus source of medium phosphorus compounds genes expression differences, and the changing of genes expression quantity as the extension of incubation time.The same incubation time, in the pH value of 5~9 NBRIP medium wj1 and wj3 strains variation trend was consistent. At 12 h, genes expression in the pH value of 5 NBRIP medium was the biggest. At 48 h, genes expression in the pH value of 9 NBRIP medium was the biggest.In summary,this study results showed the P solubilization from Ca-P was associated with a drop in the pH of the culture medium and acid secretion by phosphate solubilizing bacteria was induced by insoluble phosphate.Genes GDH was clonedsuccessfully.GDH genes expression levels of dissolved phosphorus is also affected by the type and broth medium pH.