Preliminary Study On The Intrinsic Mechanism Of Soil Component Influencing The Natural Transformation Of Bacillus Subtilis

In order to effectively assess the ecological safety of the release of genetically modified organisms into the soil environment,it is necessary to fully understand the possibility of natural transformation of soil bacteria.In this study,we chose a soil bacteria Bacillus subtilis,three kinds of common clay minerals(kaolinite,montmorillonite and goethite)and the different proportions of mineral humic acid complex as study materials,the CSF(competence and sporulation stimulating factor)pheromone,which is an important regulatory signal molecule for B.subtilis forming competence,was also introduced.The effects of three kinds of clay minerals and the interaction with CSF on the natural transformation of B.subtilis were investigated,combined with other research results that the transformation ability of the immobilized plasmid and the effect of mineral on the metabolic activity of the bacteria,the intrinsic mechanism of the mineral affecting natural transformation of B.subtilis was explained.It also preliminarily explored the effects of different proportions of mineral-humic acid complexes and the interaction with CSF on the natural transformation efficiency of B.subtilis,and analyzed the effect of humic acid on the natural transformation of B.subtilis.The main results of this study are as follows:1.Three kinds of mineral surface adsorbed plasmid pHTG and the free plasmid were transformed B.subtilis.The plasmid pHTG adsorbed on the surface of the three minerals has the ability to transform B.subtilis,and the transformation efficiency is lower than that of the free plasmid.The transforming activity of plasmids immobilized on goethite surface decreased most and the conversion efficiency of montmorillonite and kaolinite-fixed plasmids decreased the less,indicating that the goethite-binding plasmid has the lowest transformation ability.2.The effects of different types and concentration of minerals on the natural transformation of B.subtilis were studied,and the natural transformation of B.subtilis after interaction between CSF and minerals was compared.Different types and concentrations of minerals were found to have different effects on the transformation of B.subtilis.The transformation ability of bacteria from high to low is kaolinite,goethite and montmorillonite under low concentration mineral condition,but the order is goethite,montmorillonite and kaolinite under high concentration mineral conditions.The interaction of CSF with different types and concentrations of minerals has different effects on the natural transformation efficiency of B.subtilis,because minerals can affect the bioavailability of the signal molecule CSF.In this study,we think that the transformation of B.subtilis is affected by adsorption of DNA and formation of competent when the mineral concentration is low.In the presence of high concentration minerals,the effects of minerals on the transformation of B.subtilis are more influenced by the interaction between mineral and extracellular signal molecules on bacterial competence.It indicates that the interaction between soil particles and signal molecule is an important factor affecting the natural transformation of bacteria.3.The effects of the mineral humic acid complex with different proportions of three minerals and the interaction with CSF on the natural transformation of B.subtilis were studied,and the effect of humic acid on the natural transformation of B.subtilis was preliminarily analyzed.Compared with pure minerals,the combination of three minerals with humic acid can promote the natural transformation of B.subtilis in varying degrees,and the degree of promotion is influenced by the content of humic acid.The results will help us to understand the effect of soil components on the natural transformation efficiency of B.subtilis,and provide a theoretical reference for assessing the ecological risk of engineering bacteria in the environment.