Isolation And Identification Of Potassium Solubilizing Bacteria In Rhizosphere Of Robinia Pseudoacacia And Its Coinoculation Growth Promoting Effect With Arbuscular Mycorrhiza

Black locust（Robinia pseudoacacia L.）has strong resistance and adaptability.It is a pioneer tree species of soil and water conservation and one of the main artificial forest tree species of the Loess Plateau.Potassium is one of the three major nutritional elements necessary for black R.pseudoacacia and development.The Loess Plateau has serious ecological problems with soil erosion,and nutrients such as available potassium have also been lost.Most of the soil potassium is stored in potassium ore such as Yunmu and Changshi.These potassiums cannot be directly absorbed by plants.Potassium solubilizing bacteria naturally exist in nature and can decompose insoluble minerals in the soil.Arbuscular mycorrhiza fungus（AMF）can form a symbiosis with R.pseudoacacia and promote its growth.There are few studies on the effects of co-inoculation of AMF and potassium solubilizing bacteria on R.pseudoacacia.This study used a dilution plate separation method to isolate and screen potassium solubilizing bacteria from the rhizosphere soil of R.pseudoacacia in Hongjia Town,optimized its fermentation conditions;pot experiments were conducted to analyze the effects of AMF and potassium solubilizing bacteria on the growth of R.pseudoacacia seedlings;cloning potassium transporter related genes from R.pseudoacacia and analyzing the effect of AMF and potassium solubilizing bacteria on the expression of potassium transporter related genes in R.pseudoacacia.It reveals the mechanism of the influence of AMF and potassium solubilizing bacteria on the growth of R.pseudoacacia.The main results are as follows:1.Isolation,identification and optimization of fermentation conditions of rhizosphere potassium solubilizing bacteria of Robinia pseudoacaciaTwelve strains of potassium solubilizing bacteria were screened from the rhizosphere soil of R.pseudoacacia,and the rate of potassium-dissolving of each strain was between 11.99%and 25.86%.The four strains of bacteria JK2-1,JK2-10,JK2-13 and JK4 with the best potassium dissolving rate were identified as Pseudomonas by 16S r DNA,and the optimal fermentation conditions of the four strains were determined as initial p H:JK2-1 p H 7.0,JK2-10,JK2-13 and JK4 p H 8.0;Liquid content:all 10 m L;Inoculation amount:9%for JK2-1,JK2-10 and JK2-13,and 7%for JK4;Carbon source:JK2-1 and JK2-13 are sucrose,and JK2-10 and JK4 are maltose;Nitrogen source:JK2-1,JK2-10 and JK2-13 are urea,and JK4 is ammonium sulfate;Inorganic salt:JK2-1 is potassium sulfate,JK2-10 and JK2-13 are sodium phosphate,and JK4 is potassium phosphate.2.Effect of co-inoculation of potassium solubilizing bacteria and AMF on R.pseudoacaciaThe pot experiment of inoculation showed that four strains of potassium solubilizing bacteria had improved the growth of R.pseudoacacia,among which JK2-1 had the best effect.After inoculation,the fresh weight of R.pseudoacacia increased by 84.13%,the dry weight increased by 96.97%,the total potassium content of R.pseudoacacia increased by 26.25%,and the soil available potassium increased by 8.10%.Co-inoculation of 4 strains of potassium solubilizing bacteria and AMF significantly promoted the plant height,dry weight and potassium content of R.pseudoacacia,among which the plant height,dry weight,potassium content and soil available potassium content of JK2-1 and AMF co-inoculated were 45.39%,120.58%,33.63%and 12.62%higher than those of the control group,respectively.Potassium solubilizing bacteria isolated from the rhizosphere of R.pseudoacacia can decompose insoluble potassium for plant absorption and promote the growth of R.pseudoacacia together with AMF.3.Cloning and analysis of R.pseudoacacia potassium transporter geneThe Open Reading Frame Viewer（ORF）of Rp AKT1 gene is 2646 bp,and the predicted number of amino acids is 881,the molecular weight of the protein is 98870.45Da,and the theoretical isoelectric point is 6.51.The molecular formula is C₄₄₄₈H₆₉₆₃N₁₂₀₇O₁₂₈₁S₃₂.It is predicted that Rp AKT1 has 6 transmembrane regions and is a typical Shaker family transmembrane protein.The results of quantitative PCR showed that co-inoculation of AMF and potassium solubilizing bacteria could significantly up-regulate the expression of Rp AKT1,Rp SKOR and Rp HAK genes,indicating that co-inoculation of AMF and potassium solubilizing bacteria played an important role in the process of potassium absorption of R.pseudoacacia.The complementary experiment of potassium deficient yeast showed that Rp AKT1 had potassium transport function.In summary,efficient potassium solubilizing bacteria can be isolated and screened from the Loess Plateau region,and optimizing their fermentation conditions can improve the cultivation efficiency of potassium solubilizing bacteria.Inoculating potassium solubilizing bacteria can significantly increase the soil available potassium content,ultimately increasing the potassium content of Robinia Pseudoacacia plants and promoting their growth and development.AMF can form symbiosis with Robinia Pseudoacacia,and inoculation with AMF can change the root to shoot ratio of Robinia Pseudoacacia and promote its growth and development.Co-inoculation of potassium solubilizing bacteria and AMF can promote the growth and development of Robinia Pseudoacacia.Potassium solubilizing bacteria can decompose mineral potassium,and Robinia Pseudoacacia can absorb a larger range of potassium through AMF hyphae,accumulate more potassium,and significantly increase the dry and fresh weight of Robinia Pseudoacacia.In addition,inoculation with potassium solubilizing bacteria and AMF can increase the expression of related potassium transport proteins,achieving potassium absorption by improving potassium transport efficiency.This study provides a theoretical basis for the application of potassium solubilizing bacteria and AMF in locust seedling cultivation,afforestation,and sustainable forestry production in the Loess Plateau region.