Isolation Of Endophytic Actinomycetes In Rice For Cadmium Removal And Construction Of Biosensor Detection System For Heavy Metals

Recently,the rapid development of over-mining and metal smelting industry has caused serious heavy metal pollution to the environment.Heavy metal ions have strong biological toxicity,causing damage to human liver,kidney and bone.In this study,based on the detection and bioremediaton of heavy metal,endophytic strains were isolated from rice,and a biosensor for heavy metal detection was constructed to solve the problem of heavy metal pollution.Bioremediation of heavy metal pollution has the advantages of low cost and environmental protection.In this study,two strains of endophytic actinomycetes,1025-4 and 1112-10,were successfully isolated from the root of rice,and it was found that the strains jointing with plants could not only promote the growth of rice seedlings,but also have the potential to reduce heavy metal pollution.The interaction between the strains and rice seedlings was carry out to verify the growth promoting ability of the strains.Taking the untreated group as the control,the root length and stem length of rice which treated with the strain1025-4 and 1112-10 after 7 days of culture were larger than that of the control group.The root length and stem length were 102.50±11.17mm,54.00±6.10mm for the untreated group,116.33±7.97mm and 76.67±6.06mm for treated by strain 1025-4,and the 1112-10 interaction group were respectively117.83±8.61mm and 65.83±10.85mm.It indicated that the growth of rice seedlings was promoted after the interaction with the endophytic actinomycete 1025-4 and 1112-10.The cadmium MIC values of 1025-4 and 1112-10 were 0.2 mmol/L and 0.1mmol/L,respectively,indicating they has a certain resistance to cadmium.Under the condition of 0.01 mmol/L cadmium ion stress,the root length of rice seedling treated by 1025-4and 1112-10 were increased by 8.33±1.93 mm and 10.5±2.43 mm respectively,comparing with the untreated group after 7 days cultivation.The growth of rice seedling was better than that of the control group,indicating that the interaction of rice and the endophytic strains 1025-4 and 1112-10 could improve the resistance of rice seedlings to the cadmium stress.It will provide the microbial resources for bioremediation of heavy metal pollution.A biosensor system for the detection of heavy metal copper,nickel and cadmium was constructed,which was mainly composed of the fluorescent protein m Apple and the metal binding protein D6A3.The fluorescence intensity of the sensing protein m Apple-D6A3 decreased in the presence of heavy metal ions such as cadmium,nickel and copper.It was proved by isothermal titration quantitative thermometry?ITC?that the sensing protein m Appple-D6A3 is bound to copper,nickel and cadmium ions,which may be due to the conformational change of the protein.The fluorescence intensity of the sensing protein m Apple-D6A3 is greatly affected by the p H.The sensor protein will be destroyed under acidic conditions,while it is relatively stable under weak alkaline conditions.p H 9.0 was used in this study.The ability of the sensing protein m Apple-D6A3 to detect copper,nickel and cadmium ions was verified.It was found that there was a strong linear relationship between the fluorescence intensity of the protein and the ion concentration,and the correlation coefficients were 0.98(Cu²⁺),0.94(Ni²⁺)and 0.99(Cd²⁺),indicating that the sensing protein m Apple-D6A3 had the ability to detect copper,nickel and cadmium ions of heavy metals.In order to verify the practical application ability of the biosensor,copper,nickel and cadmium ions in water and medium were detected,and the accuracy of the results was more than 85%.The rapid detection and bioremediation are of great significance to the treatment of heavy metal pollution.In this study,endophytic strains with bioremediation potential of heavy metal contamination were isolated from rice,and a fast detection method of heavy metal ions using a biosensors system based on fluorescent proteins was proposed to provide a reference for the treatment of heavy metal pollution in the environment.