44 results about "Microcystin-LR" patented technology

The invention provides a process for utilizing three abandoned proteins and thoroughly-decomposed cow dung to produce a growth-promoting bio-organic fertilizer and a product thereof and belongs to agricultural intensive production technologies. A used formula comprises, by dry weight, 75% of thoroughly-decomposed cow dung ordinary manure, 5% of alga ooze, 10% of abandoned feather puffing powder and 10% of rapeseed meal. A used strain is a growth-promoting bacillus amyloliquefaciens SQR-9. The process comprises mixing the raw materials according to the optimum formula, adding 5% of growth-promoting bio-control bacteria by volume mass, performing strip stack type fermentation, performing overturning once each day, and fermenting for 6-7d. The number of beneficial viable bacteria in the produced fertilizer is lager than 2*10<8>CFU / g by dry weight (similarly hereinafter), the content of total nitrogen is 2-3%, the content of total NPK nutrient is 6-8%, the content of organic matter is larger than 45%, and the content of microcystin LR and the content of microcystin RR are both lower than 4ug / kg. The growth-promoting bio-organic fertilizer is rich in beneficial amino acid, low in production cost and high in growth-promoting effect and can be applied to food crops and vegetables and the like.

The invention relates to a method for detecting microcystin-LR by an immunochromatographic test strip and belongs to the technical field of immunoassay chemistry. The method comprises the following steps: preparing Au NCs with a hollow structure by a one-step preparation method, marking Au NCs on Anti MC-LR to prepare a gold nanocage-microcystin antibody conjugate, and respectively spraying BSA (bovine serum albumin)-coupled microcystin-LR and IgG to a nitrocellulose membrane for being used as a detection line and a control line to prepare an immunochromatographic strip. The microcystin-LR in a test sample is analyzed in a quantitative manner by reading the gray-scale value of the gold nanocage-microcystin antibody conjugate which is retained on the detection line through a competitive immunochromatographic method. The shortcoming that gold sol is not easy to preserve in the technology of the test strips can be overcome; furthermore, the operation is simple, and the time required for modifying an antibody is relatively short.

The invention relates to a visible light driving Cu2O / PPy composite material based microcystins-LR molecular imprinting photo-electrochemical sensor and preparation and application thereof. The visible light driving Cu2O / PPy composite material based microcystins-LR molecular imprinting photo-electrochemical sensor is characterized in that ITO conducting glass is used as a substrate, and a molecular imprinting film is deposited on the substrate through an electrochemical technology, wherein the molecular imprinting is prepared by compositing a photoelectric material which is a Cu2O semiconductor that can be absorbed within the visible light range and polypyrrole containing microcystins-LR vacancy; the visible light is treated as an excitation light source; the concentration of microcystins-LR is detected by i-t technology through a CHI660e workstation. Compared with the prior art, the microcystins-LR molecular imprinting photo-electrochemical sensor has the advantages of being sensitive to photo-electrochemical response, wide in detection range, capable of detecting within the concentration range of 1.0ng.L<-1> to 10.0 mu g.L<-1>, and high in interference resistance. The preparation method is simple, quick, low in cost, and applicable to quick online detection of water.

The invention relates to a method for screening single chain antibodies of Microcystin-LR and verification thereof. The method comprises the following steps: carrying out two rounds of affinity screening on the biotinylated Microcystin-LR in a human source synthetic antibody library by using an avidin labeled magnetic bead and a negative screening method; extracting total plasmid DNA from phage colonies produced in the second round, carrying out enzyme digestion with enzyme Sfi I, recycling gel to obtain single chain antibody genes, connecting the single chain antibody genes with soluble expression carrier pAK100CL which is processed by enzyme digestion in the same way, and electrically transforming the connected carrier into colibacillus XL1-Blue to obtain soluble expression single chain antibodies; and verifying the soluble expression single chain antibodies by using a competitive time-resolved fluorescence immune analytical method. The invention has the advantage of quick, simple and convenient screening, and can well expose the Microcystin-LR three-dimensional structure into the incubation system. The verification on the screening result has the advantages of high detection signal and strong anti-interference capacity against stroma.

Preparation and use methods of a microcystin-LR (MC-LR) monoclonal antibody immunoaffinity column (IAC) belong to the technical field of immunoaffinity chromatography and MC detection. The IAC is prepared by fixing MC-LR monoclonal antibody in the column, and can acquire the accurate contents of MC-LR and MC-RR through solvent extraction of cyanobacteria sample, enrichment of a solid-phase extraction column, IAC purification and quantitative determination by liquid chromatography-mass spectrometry. The determination can be interfered seriously by impurities to influence the result accuracy by only using SPE purification without IAC purification. The methods can be used for MC detection in water sample, other algae and similar biological samples.

The invention discloses a fluorescent probe for detecting microcystic toxin-LR in water, which belongs to the field of molecular biology and microbiology. According to the fluorescent probe, MC-LR is taken as a target, a dodecapeptide library randomly shown on the surface of a bacteriophage is used for selecting a recombination bacteriophage which is combined with the MC-LR in a specific manner, and the single-stranded DNA (Deoxyribonucleic Acid) of the recombination bacteriophage is extracted and recombined for sequencing and sequence alignment, thereby obtaining an MC-LR specific affinity ligand sequence: His-Phe-Phe-Lys-Trp-His-Thr-Arg-Thr-Asn-Asp-Gln, and subsequently, obtaining the specifically combined fluorescent probe FITC-Acp-His-Phe-Phe-Lys-Trp-His-Thr-Arg-Thr-Asn-Asp-Gln through solid-phase polypeptide synthesis and fluorescent marks. The fluorescent probe has the beneficial effects that firstly, a completely new tool is provided for the specific identification and content detection of the MC-LR; and secondly, the fluorescent ligand probe can be used for qualitatively identifying the MC-LR and quantitatively detecting the content of the MC-LR in a sample.

The invention discloses a method for adsorbing and removing phycobiliprotein and microcystin-LR by modified attapulgite compounded by organic polymer, comprising the steps of compounding the attapulgite and chitosan powder according to the ratio of 50-100:0.5-2 by mass, adding the compounded mixture into waste water containing phycobiliprotein and microcystin-LR according to the ratio of 0.1-2g / L, stirring, standing, and spooning or filtering to remove the floc. The invention has the characteristics of low cost, high removal efficiency and environmental friendliness, and has larger application value and market prospect.

The invention discloses a method for detecting microcystin-LR by self-assembly based on the end face of a gold nano-rod, belonging to the technical field of chemical immunoassay. In the invention, the end face of the gold nano-rod is respectively coupled with microcystin coating antigen and anti-microcystin antibody to form a probe of the gold nano-rod; and the synthesized probe of the gold nano-rod performs immunological reaction with the microcystin-LR so as to achieve the detection of the microcystin-LR through the change of the grain diameter of a nano-chain which is formed by the antigen antibody reaction of the end face of the gold nano-rod. The invention performs reaction in a liquid environment, only one step of reaction is needed without washing step, which simplifies the condition of the reaction, reduces labour intensity and improves the detection sensitivity.

The invention discloses an alga lysing / algal toxin degradation double-effect engineering strain Y1 and a construction method thereof, belonging to the field of microbiology for treatment of blue-green algae. Fusion is performed on parent strains with alga lysing and algal toxin degradation functions by utilizing a protoplast fusion technology to construct the double-effect engineering strain with excellent traits of parents, namely Bacillussphaericus with the collection number of CGMCCNO. 7519. The alga lysing and the derived algal toxin pollution problem are synchronously solved. The protoplast fusion rate can achieve 31.97% under optimal conditions, and the double-effect engineering strain has certain guide effects on solving the problems of blue-green alga bloom and the derived MC-LR (microcystin-LR) pollution and constructing the engineering strain integrating the MC-LR degradation function and the alga lysing property into a whole by performing a protoplast fusion technology, thereby providing reference for achieving the effect on treating both the root course and the symptoms for solving the large-area blue-green alga bloom problem.

The invention relates to a preparation method of an enzyme-free immuno-sensor for detecting microcystin-LR (MC-LR) and a detection method thereof, and belongs to the technical field of immuno-analytical chemistry. According to the method, carboxylated carbon nano fiber and polyethylene glycol are assembled on the surface layer of a glass-carbon electrode to construct a high density MC-LR antigen sensing interface; gold nano particles are synthesized, a gold nano particle probe is prepared, the indirect competition immunoassay principle is adopted, embedded MC-LR antigen and MC-LR target antigen compete for MC-LR antibody to form an immune complex, the gold nano particle probe reacts with the immune complex; the gold nano particles bonded on the surface of the electrode are oxidized to generate AuCl<4->, the value of the peak current of the reduction peak of AuCl<4-> can be measured by an electrochemical method, and finally the MC-LR is detected through the value of the peak current. The provided method can online detect MC-LR high sensitively, the purification treatment of a water sample is not needed, the detection is simple and rapid, the detection system does not need any enzyme, and thus the detection cost is reduced.

The invention provides a dual-effect engineering bacterium Y7 capable of lysing algae and degrading algal toxin and a construction method thereof, and belongs to the microbiological field of blue-green algae treatment. The dual-effect engineering bacterium Y7 capable of lysing algae and degrading microcystin-LR (MC-LR) is prepared based on a protoplast fusion technology in which the protoplast fusion rate can be up to 28.17% under optimal conditions. The dual-effect engineering bacterium Y7 provided by the invention is used for solving the problems of the outbreak of blue-green algae and the pollution of derived MC-LR, plays a guiding role in constructing a engineering bacterium integrating an MC-LR degradation function and algae-lysing property by virtue of an improved protoplast fusion technique and practical application and provides reference for solving the problem large-scale blue-green algae blooms fundamentally.

Preparation and use methods of a microcystin-LR (MC-LR) polyclonal antibody immunoaffinity column (IAC) belong to the technical field of immunoaffinity chromatography and MC detection. The IAC is prepared by fixing MC-LR polyclonal antibody in the column, so that MC-LR in the sample solution can be adsorbed by the IAC and can be enriched and purified after elution, then the quantitative and qualitative analysis can be carried out by HPLC. As the pre-processing process, the IAC enrichment is superior to that of the prior solid-phase extraction (SPE) method. HPLC test shows that MC-LR peak of the MC-LR polyclonal antibody can be separated from peaks of other impurities by IAC so as not to influence the detection result. However, the SPE method is difficult to distinguish the MC-LR peak due to the presence of so many impurities. The MC-LR polyclonal antibody can be specifically combined with the MC-LR, so that IAC has excellent specificity to remove the majority of interferents in one step. The SPE has no specificity and cannot remove the interferents so as to influence the final detection result.

The invention discloses a hybridoma cell strain YNEMC3 and a monoclonal antibody generated thereby. The preservation number of the murine microcystin LR monoclonal antibody hybridoma cell strain YNEMC3 in China Center for Type Culture Collection is CCTCC NO:C201298. The results of experiments about the application of the monoclonal antibody in the murine microcystin LR detection show that the monoclonal antibody is specific to the murine microcystin LR and greatly eliminates the interferences of other microcystins in order to obtain an accurate determination result.

The invention relates to a method for preparing microcystin-LR. In the method, blue-green algae obtained from Dianchi in Yunnan are taken as raw materials. The method comprises the following steps of: performing extraction by using 50 to 90 percent (v / v) aqueous solution of acetonitrile to obtain crude microcystin, and removing impurities by membrane filtration; performing primary purification and enrichment by adopting solid phase extraction; and performing separation and purification by semi-preparative high performance liquid chromatography. In the pre-treatment step of the method, most of the impurities are removed, so the purification difficulty of the high performance liquid chromatography is reduced; the method has the advantages of simplicity, easy operation and relatively less microcystin loss; and the purity of the finally obtained microcystin-LR is 91.35 percent which is relatively higher.

The invention comprises a photoautotrophic organism, generally having simpler nutritional requirements than heterotrophic organisms, utilized as a chassis for the heterologous expression and function of enzymes, or derivatives of said enzymes, that show activity toward the degradation / detoxification of toxins known to be associated with and specific to harmful algal blooms. As an example, a cyanobacterial strain (Synechocystis sp. PCC 6803) modified to express Sphingomonas sp. USTB-05 MlrA enzyme functionality, showing the capability of degrading microcystins (results shown here) and nodularins, is presented. Under modelled natural conditions, results indicate that heterologous enzymatic activity against microcystin-LR is more stable over time when utilizing a photoautotrophic chassis in comparison to use of a heterotrophic bacterial strain. In addition, both the viability and cell density of the photoautotrophic host is maintained for a significantly longer period of time, compared to a heterotrophic host.

The invention discloses a method for detecting microcystin-LR in water through high performance liquid fluorescence chromatography, in particular relates to the method for detecting the microcystin-LRin the water through the high performance liquid fluorescence chromatography and belongs to the field of a water environment monitoring technology or an agricultural monitoring technology. Accordingto the method, a sample is filtered, enriched, eluted and concentrated to obtain a constant volume based on the characteristics that fluorescent light can be stimulated by the microcystin-LR under theultraviolet irradiation; rapid, accurate and efficient detection of the microcystin-LR in a water body is realized by adopting the high performance liquid fluorescence chromatography; the method hasguiding significance on monitoring, management and protection of a water body pollution condition.

The invention provides a dual-effect engineering bacterium Y7 capable of lysing algae and degrading algal toxin and a construction method thereof, and belongs to the microbiological field of blue-green algae treatment. The dual-effect engineering bacterium Y7 capable of lysing algae and degrading microcystin-LR (MC-LR) is prepared based on a protoplast fusion technology in which the protoplast fusion rate can be up to 28.17% under optimal conditions. The dual-effect engineering bacterium Y7 provided by the invention is used for solving the problems of the outbreak of blue-green algae and the pollution of derived MC-LR, plays a guiding role in constructing a engineering bacterium integrating an MC-LR degradation function and algae-lysing property by virtue of an improved protoplast fusion technique and practical application and provides reference for solving the problem large-scale blue-green algae blooms fundamentally.

The invention provides preparation and application of a quantitative and rapid detection biosensor of microcystic toxin-LR, and belongs to the biotechnology field. The detection sensor is an impedance type electrochemical sensor, and consists of a CHI760C type electrochemical work station and a three-electrode system in which an immunity electrode is taken as a working electrode, a saturated calomel electrode is taken as a reference electrode and a platinum wire electrode is taken as a counter electrode; L-cysteine, colloidal gold and microcystic toxin antibody compound are respectively enveloped on a surface of a bare mitallic electrode to be taken as the working electrode; a change of electric potential difference between the reference electrode and the working electrode takes place when the microcystic toxin antibody is combined with the antigen (MC-LR) in sample solution, and is subject to a quantitative relation with the concentration of the antigen in the sample, then the detection of the microcystic toxin is carried out; the biosensor for rapidly, specifically and sensitively detecting microcystic toxin antigens is successfully established; the detection linear range is 0.05-300ng / mL, and the detection limit is 1.82*10<-2>ng / mL. The biosensor can perform real-time monitoring on natural lake water, and can also perform real-time monitoring on preparation process of drinking water in water supply works.

The invention relates to a method for removing Microcystin-LR in water body by using bromides, which is characterized by comprising the following steps: preparing a 1,3-Dibromo-5,5-dimethylhydantoin solution with the concentration of 100-10000 mg / L, and spraying the 1,3-Dibromo-5,5-dimethylhydantoin solution to a Microcystin-LR polluted water area by using a spraying machine, wherein the spray amount of the 1,3-Dibromo-5,5-dimethylhydantoin solution is 100-300 ml / m<2>; and 1-60 minutes after spraying the 1,3-Dibromo-5,5-dimethylhydantoin solution to the water area, the content of effective bromine in the water area is 0.1-100 mg / L. The invention has strong effect on removing the Microcystin-LR by utilizing the reaction activity of 1,3-Dibromo-5,5-dimethylhydantoin, and can not produce secondary pollution, or teratogenetic, cancerigenic and mutagenic products. The method has the advantages of high removal rate, high toxicological safety, low investment and strong operability, and can be used for removing the Microcystin-LR in the water body.

The invention discloses a microcystin-LR enzyme-linked immune detection kit, and belongs to the technical field of enzyme-linked immune analysis. The kit comprises a box, an enzyme label plate and a reagent, the enzyme label plate and the reagent are arranged in the box, and microcystin-LR is detected by an ELISA (enzyme-linked immunosorbent assay) direct competition method. The kit is characterized in that the enzyme label plate wraps microcystin-LR antibodies, and the reagent comprises microcystin-LR horse radish peroxidase labeled antigen solution, microcystin-LR standard solution, condensed scrubbing solution, substrate developing solution and reaction stop solution. The enzyme-linked immune detection kit has the advantages that the kit is simple, convenient, rapid, sensitive, accurate and low in cost, the sensitivity of the kit can reach 0.05ng / mL, and the kit is particularly application to residue detection of microcystin-LR in water and algae food.

The invention discloses a method for detecting microcystin-LR in a high sensitivity mode by applying dynamic light scatting, which belongs to the technical field of immunoassay chemistry. A gold nanoparticle synthesized by the method couples with a coating antigen of microcystin to obtain a gold ligated coating antigen, and is connected with an antibody against the microcystin to obtain a gold ligated antibody, the antibody and the gold nanoparticle are used for an immune reaction, trisodium citrate is utilized to reduce a dimmer, an oligomer, and a coagulation formed by the reaction of the gold ligated antibody, the gold ligated coating antigen and the microcystin in a liquid environment, and the content of the microcystin is measured by measuring particle diameter distribution in the liquid environment through dynamic light scattering. The method reacts in liquid environments only, does not need the step of washing, only needs a one-step reaction, simplifies reaction conditions, improves the sensitivity of the detection, and reaches the leading level in the world.

The invention relates to preparation and application of a microcystin-LR quantitative and rapid detection sensor, and belongs to the technical field of biology. The detection sensor is an impedance type electrochemical sensor and consists of a CHI760C type electrochemical working station and three-electrode system, wherein the three-electrode system comprises an immunoelectrode serving as a working electrode, a saturated calomel electrode serving as a reference electrode and a platinum wire electrode serving as a counter electrode; an L-cysteine, colloidal gold and microcystin antibody composition is wrapped on the surface of a bare gold electrode to prepare the working electrode; when a microcystin antibody is combined with an antigen (MC-LR) in sample liquid, a change of a potential difference between the reference electrode and the working electrode is caused; the change and the antigen concentration of a sample are in a quantitative relation, so as to carry out detection of microcystins; a fast, specific and sensitive biosensor for detecting a microcystin antigen is successfully constructed; the detection linear range is 0.05 to 300 ng / mL, and the detection limit is 1.82*10<-2> ng / mL. The microcystin-LR quantitative and rapid detection sensor can monitor natural lake water in real time and also monitor a preparation process of drinking water for a water plant in real time.

The invention discloses an alga lysing / algal toxin degradation double-effect engineering strain Y1 and a construction method thereof, belonging to the field of microbiology for treatment of blue-green algae. Fusion is performed on parent strains with alga lysing and algal toxin degradation functions by utilizing a protoplast fusion technology to construct the double-effect engineering strain with excellent traits of parents, namely Bacillussphaericus with the collection number of CGMCCNO. 7519. The alga lysing and the derived algal toxin pollution problem are synchronously solved. The protoplast fusion rate can achieve 31.97% under optimal conditions, and the double-effect engineering strain has certain guide effects on solving the problems of blue-green alga bloom and the derived MC-LR (microcystin-LR) pollution and constructing the engineering strain integrating the MC-LR degradation function and the alga lysing property into a whole by performing a protoplast fusion technology, thereby providing reference for achieving the effect on treating both the root course and the symptoms for solving the large-area blue-green alga bloom problem.

The invention discloses a method for evaluating toxicity of Microcystin in a real water environment. The method comprises the following steps: preparing coexistence liquid of degrading strains and Microcystin-LR (MC-LR) and constructing a contamination sample with the form close to that of the MC-LR in a real water environment; then infecting human hepatoma carcinoma cells, and verifying toxicityof MC-LR under the condition of coexistence of the degrading strains by using an MTT method for detecting cell metabolic activity. According to the invention, the state of MC-LR in coexistence with the degrading strain in real water environments such as lakes is simulated, and the Limitation of toxicity research on single MC-LR is made up for, the toxicity of MC-LR is better evaluated, and the application prospect in the industries of aquaculture, environmental governance and the like is good. The method has universal applicability to various Microcystin isomers and is wide in application range. According to the method, the degrading strains is irradiated by an N+ ion beam injection technology to obtain irradiated degrading strains, the irradiated degrading strains are compared with the degrading strains before irradiation in the aspect of degrading effect on MC-LR, and then the feasibility of the method is verified.

The invention discloses a method for detecting microcystin-LR by self-assembly based on the end face of a gold nano-rod, belonging to the technical field of chemical immunoassay. In the invention, the end face of the gold nano-rod is respectively coupled with microcystin coating antigen and anti-microcystin antibody to form a probe of the gold nano-rod; and the synthesized probe of the gold nano-rod performs immunological reaction with the microcystin-LR so as to achieve the detection of the microcystin-LR through the change of the grain diameter of a nano-chain which is formed by the antigenantibody reaction of the end face of the gold nano-rod. The invention performs reaction in a liquid environment, only one step of reaction is needed without washing step, which simplifies the condition of the reaction, reduces labour intensity and improves the detection sensitivity.

The invention relates to a microcystic toxin-LR electrochemical detection method with amplified graphene signals. The method comprises the following steps: (1) dropwise adding a mixed solution with a ratio of an endonuclease reaction buffer solution: a graphene-aptamer compound: a microcystic toxin-LR solution to be detected: an endonuclease solution of (9-11):(4-6):(2-4):(1-3) on the surface of an MCH / Au NPs / Au electrode; (2) culturing the MCH / Au NPs / Au electrode for 1 to 2 hours at a temperature of 25 to 35 DEG C, slightly washing the electrode surface by ethanol and ultrapure water to remove weakly combined graphene, drying the electrode in nitrogen gas, taking the electrode out, carrying out electrochemical detection to measure the concentration of the microcystic toxin-LR solution. According to the provided method, a high sensitive electrochemical analysis technology and an aptamer with a high specific recognition performance are effectively combined to detect trace microcystic toxin-LR in the environment, the detection is highly sensitive and highly specific, moreover, the instruments are cheap and portable, the method is simple and easy to perform, and the results can be obtained quickly.

The invention provides a method for detecting MC-LR (microcystin LR) in a sample solution. The method comprises the steps as follows: a nucleic acid aptamer capable of specifically binding the MC-LR is fixed on a micro-cantilever array; the micro-cantilever array fixed with the nucleic acid aptamer is contacted with the sample solution; bending deformation produced after the micro-cantilever arrayfixed with the nucleic acid aptamer is contacted with the sample solution is detected. The invention further provides the micro-cantilever array and a system for detecting the MC-LR in the sample solution.

The invention relates to a method for detecting microcystin-LR through a palladium-gold alloy nanoscale cage immune chromatography test strip, and belongs to the technical field of immunity chromatography chemistry. The method comprises the following steps: preparing a Pd-Au bimetal nanoscale cage with a hollow structure by taking Pd nano-particles as seed crystals; spraying a monoclonal antibody for labeling the microcystin-LR to a combined pad; spraying BSA (Bovine Serum Albumin) coupling microcystin-LR on a detection line; spraying goat anti-rat IgG on a control line to prepare the immune chromatography test strip for fast detecting the microcystin-LR in a water body; and carrying out quantitative analysis on the microcystin-LR contained in a test sample by a competition immune chromatography and by reading the grey value of gold nanoscale cage-microcystin antibody mixture remained on the detection line. According to the method, because the adhesion of sulfhydryl and amido in a protein molecule of a Pd-Au alloy is higher than the binding force of a single-metal gold nanoscale cage by virtue of the synergistic action of two metal elements in the alloy, a microcystin antibody is easier to label by that Pd-Au alloy nanoscale cage than by the gold nanoscale cage, and the formed Pd-Au alloy nanoscale cage-microcystin antibody mixture is more stable.

The invention relates to the technical field of water treatment microorganisms, and in particular relates to preparation and regeneration method of an algal toxin degrading bacterium protoplast. The bacterium is separated from blue-green algae dehydration filtrate and is identified to be a capitatus lysine bacillus and is preserved in the China general microbiological culture collection center, the preservation number is China general microbiological culture collection center (CGMCC) NO.6107, and the preservation name is microcystin-LR (MC-LR) degrading bacterium R3. The preparation and regeneration method mainly comprises bacterium seed culture solution preparation, hypha suspension preparation, cell wall enzymolysis, protoplast suspension preparation and protoplast regeneration. The protoplast preparation and regeneration are adopted, the preparation rate of the protoplast for degrading the capitatus lysine bacillus R3 of the MC-LR can reach 85.10%, and the regeneration rate can reach 38.40%; and moreover, the preparation and regeneration processes are low in equipment requirement and are simple to operate, the enzymolysis time and regeneration period are short, and the subsequent fusant preparation and engineering bacteria structure of R3 are benefited.