| With the widespread use of chemical fertilizers,some farmLand of China has been excessive fertilized,and it caused series of ecological problems.As important vegetable cultivation base,the Pearl River Delta region also has these problems.Study on the changes of soil microbial community structure and function in long-term excessive fertilization can provide some guidance for proper fertilization of farmLand.In this study,we focused on alkaline phosphatase,firstly did research about farmLand soils in Guangzhou area by combining physical and chemical indicators detection and high-throughput sequencing.A strain organophosphorus degrading bacteria GN1 was isolated from the farm Land soil,and cultured in inorganic phosphorus medium in long-term to study its alkaline phosphatase genes' expression and the enzyme activity under long-term inorganic phosphorus stress.At the same time,representative farmLand soil was taken into simulated fertilization to study the transcription and expression of alkaline phosphatase and the change of enzyme activity during phosphate fertilization.The main conclusions of this study are as follows:Six soil samples were collected from Panyu,Haizhu,Tianhe,Liwan and Conghua in Guangzhou.The physical and chemical indicators of samples were detected and 16 S rDNA,18 S rDNA,alkaline phosphatase gene phoD and phoX soil were taken into high-throughput sequencing.The results showed that the content of inorganic phosphorus and ammonia of farm Land soils in Guangzhou is low.Results of high-throughput sequencing showed that the microbial diversity of farm Land soil in Guangzhou was high,and the dominant phyla of bacteria community were Proteobacteri,the dominant phyla of fungal community was Ascomycota.pho D and phoX genes containing microorganisms have high homology with farm Land soil uncultured bacterium.Meanwhile,? diversity analysis of high-throughput sequencing showed that the regional differences of soil microbial diversity in Guangzhou was obivious,and the similarity between different samples was low.In the simulated fertilization experiment,the results showed that the presence of inorganic phosphorus in soil could inhibit the activity of soil microbes alkaline phosphatase in the process of fertilization,and the presence of organic phosphorus in soil would enhance the activity of soil microbial alkaline phosphatase.However,experimental results at the level of transcriptional expression of the gene showed that the expression of alkaline phosphatase in the soil was raised by the addition of either inorganic phosphorus or organic phosphorus.Combined with the results of CCA analysis in high throughput sequencing,We inferred that the cause to above phenomenon is mainly due to the unique characteristics of farmLand soil.Isolated organophosphorus degrading bacteria GN1 growed well in different phosphorus medium,but showed different enzyme activity.The result of genome sequencing showed that GN1 has low similar with other acinetobacter species that had been found,and there were rarely reported.The transcriptome sequencing of GN1 cultured with different phosphorus showed that the gene expression of GN1 was really different in different phosphorus medium.When GN1 was cultured in organic phosphorus medium,a large number of genes which include phosphorus metabolism related genes were up-regulated expression.The results of long-term inorganic phosphorus culture showed that activity of alkaline phosphatase of GN1 continuously increased during 500 days' inorganic phosphorus culture.The results of transcriptome sequencing also showed that a large number of GN1 genes including alkaline phosphatase genes were up-regulated expression during continuous culture.It could be inferred that alkaline phosphatase gene of soil microbes and its enzyme activity will be up-regulated under the constant stimulation of inorganic phosphorus for a certain period.In this experiment,because the experimental time is not long enough,we hadn't observed mutation or loss of phosphate metabolism related genes. |
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