Study On The Biosynthesis Of Near-infrared Ag₂Se Quantum Dots In Escherichia Coli

Due to their good optoelectronic properties,Quantum Dots have a wide range of applications in many fields such as in vivo imaging,drug transportation,biochips and optoelectronic devices.However,in biomedical applications,compared with visible fluorescent quantum dots,near-infrared quantum dots have stronger tissue penetration and are less affected by tissue autofluorescence,so they are ideal for deep tissue imaging in living bodies.Silver-based near-infrared quantum dots have been extensively studied in recent years due to their narrow band gap and low toxicity.Among them,Ag₂Se quantum dots,as a new kind of quantum dots,have the advantages with small size,low toxicity and good near-infrared optical properties.Ag₂Se quantum dots have stronger biocompatibility than other near-infrared quantum dots containing Cd,Pb and Hg.The synthesis methods of Ag₂Se quantum dots mainly include chemistry and biology.Compared with traditional chemical synthesis methods that require high temperature and pressure in a synthetic environment involving toxic,flammable and explosive chemical reagents,there are still fewer studies on biosynthesis methods with green environmental advantages.There have been many reports showing that the microbial cells can be used to synthesize some types of nanoparticles.Prokaryotes generally grow faster,have higher tolerance,and faster metabolism and detoxification processes,which are different from eukaryotes.Therefore,prokaryotic cells have broader research and application prospects in the biosynthesis of nanomaterials such as quantum dots.In this study,the prokaryotic organism Escherichia coli was used as the experimental material,and the"time-space coupling"strategy(the coupling of two originally unrelated metabolic pathways in time and space)as a guide,the following research content was carried out:1.First,the study of biosynthesis of near-infrared Ag₂Se quantum dots in E.coli BW25113 were carried out.Using the"time-space coupling"strategy,by measuring the fluorescence spectra of cells,it was found that E.coli BW25113 could synthesize near-infrared quantum dots.In order to further determine the synthesized product,the near-infrared quantum dots synthesized in the cell were characterized by transmission electron microscopy(TEM),energy dispersive X-ray spectroscopy(EDX)and photoluminescence(PL)emission spectroscopy.The results showed that the product was near-infrared Ag₂Se quantum dots,which was spherical particles with small particle size and uniform distribution.The statistical average particle size was 2.9�0.7nm,and the lattice spacing was 0.225 nm.Spectroscopy showed that the biosynthetic Ag₂Se quantum dots had an absorption peak at 415 nm and the highest emission peak at 920 nm,which belonged to the first near-infrared window and had good fluorescence stability.The study compared the effects of silver nitrate concentration and synthesis time on the synthesis of Ag₂Se quantum dots in E.coli.The fluorescence intensity of intracellular quantum dots was used as an indicator to detect the ability of biosynthesis of quantum dots.It was found that the synthesis effect of quantum dots was better when the concentration of silver nitrate was 1 m M,the synthesis time was at3 h.By measuring the toxicity of quantum dots to Vero cells and in vivo imaging in mice,the biosynthetic Ag₂Se quantum dots had good biocompatibility,and their fluorescence could penetrate the body of nude mice,which could be used for further in vivo imaging applications.2.From the perspective of quantum dot biosynthesis mechanism,the synthesis ability of Ag₂Se quantum dots was improved.In order to verify whether the selenium precursor had an effect on the synthesis of Ag₂Se quantum dots,firstly,the important gene cys E in the synthesis pathway of selenium precursor was knocked out,it was found that the ability of E.coli to synthesize quantum dots was significantly reduced after the deletion of the cys E gene.After overexpression,the synthesis ability could be improved.It showed that selenocysteine in E.coli was the precursor substance of quantum dot synthesis and participated in the synthesis of Ag₂Se quantum dots.Combining the selenium metabolism mechanism of E.coli,knocking out the met A gene in the selenium metabolism decomposition pathway increased the content of intracellular selenoamino acids and successfully enhanced the biosynthesis ability of Ag₂Se quantum dots in E.coli.In summary,this study has confirmed that E.coli BW25113 cells could synthesize near-infrared Ag₂Se quantum dots,and the biosynthetic Ag₂Se quantum dots had good biocompatibility and could be used for further research in biomedical applications.Selenocysteine is involved in the synthesis of E.coli Ag₂Se quantum dots,and its content could be increased through genetic modification to improve the ability of cells to synthesize Ag₂Se quantum dots.This research had opened up a new method and provided reference ideas for the preparation of near-infrared quantum dots by prokaryotes.