| Phosphorus(P) is vital for a large number of crop nutrients,but soil phosphorus is mostly insoluble so that it is difficult to be absorbed by plants. In this study, two efficient phosphorus-solubilizing Bacillus strains L1 and L2 was isolated from soil though the calcium phosphate broth enrichment culture methods. The result of maize pot and field experiments shown the improvement of strains for effectiveness of soil phosphorus, phosphorus content of plants and plant biomass. And because of the green fluorescent protein labeling method, we studied the when and where the strain L1 colonized in maize root surface. The main results are as follows:(1) Eight P-solubilizing bacteria were isolate. After culturing in 1% calcium phosphate liquid medium for 72 hours under 30 oC, the soluble P in each culture solution ranged from 73.72 mg·L-1 to 392.56 mg·L-1, wherein the P-solubilizing capacity of strain L1, L2, L7 were outstanding beyond others and the soluble P of them were 392.56 mg·L-1, 296.77 mg·L-1, 353.59 mg·L-1. To this end, by the further analysis of colony morphology, physiology and biochemistry 16 S r DNA sequencing feature for these three, we preliminarily identified that L1 and L7 was Bacillus megaterium, L2 was Bacillus aryabhattai.(2) The maize pot experiment result in June 2015 shown that, in the 25 th day, the soil phosphorus content of treatment groups L1 and L2 were increased by 30% and 21% compared with CK. The date in September 2015 maize pot experiment result show that in the first 15 days and 25 days for each strain significantly increased the effectiveness of soil phosphorus, especially the soil phosphorus content of L1 treatment have a increase of 46% than CK and increased by 32% compared with L2 at the day 25.(3) Both two microbial agents could significantly improve phosphorus accumulation in maize plant. In the first maize pot experiment, compared with control, treatment L1 respectively improved phosphorus accumulation in plant by 22.41%. Under the condition of pot experiment, treatment group L1 had an advantage over L2 in promoting the plant to uptake phosphorus.The increasing amount of treatment L1 and L2 in the second maize pot experiment was 34.04%, 27.71%, while the data in maize field trails was 55.95%, 57.63%.(4) Both two microbial agents could significantly increase plant biomass. Compared with control, treatment L1 and L2 increased dry weight of maize seedling by 18.78%, 18.62%, and increased dry weight of maize root by14.84%, 4.69%. Under the condition of field trails. The increasing amount of dry weight of maize seedling were 75.28%, 60.75%, while the increased dry weight of maize root were 100.70%, 99.82%. The increased yield of each were 9.56%, 8.09%.These results suggested that the P-solubilizing capacity of strain L1 is much more effective than L2, and the same of effectiveness of soil phosphorus, plant phosphorus accumulation and plant biomass. All the results shown the potential of L1 for improving the effectiveness and improving the soil phosphorus and maize phosphorus nutrient.For this reason, by using green fluorescent protein(GFP) labeling method,a high high fluorescence intensity, good stability strain GFP-labeled L1 was constructed through molecular cloning techniques. The gfp-L1 has high fluorescence intensity, and after a continuous culturing for 7days, the GFP plasmid stability is still as high as 87%, indicating that the strain gfp-L1 could be used build mechanisms of maize roots colonization. 2 days after maize germination, gfp-L1 were incubated t the maize. 24 h later, the green fluorescent of gfp-L1 were observed on the surface and inner maize root, which confirmed the strain L1 has been successfully colonized inside of maize root hair cells.Taken together, these results shown that strain L1 has a great potential in improving the effectiveness of soil phosphorus and phosphorus nutrient of maize. It has great potential for bio-fertilizers and being expected to play a catalytic role in the sustainable development of agriculture. At the same time under laboratory conditions, it confirms that L1 successfully colonized maize root hair surface and inside cells, which could be the basis for further research of L1 in situ soil and plant rhizosphere colonization mechanism. |
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