25 results about "Antitoxin" patented technology

The invention provides a production strain of an antibiotic-free plasmid, the production strain is a gene editing strain of a PIR strain and is named as PIR1-WN:: 0636 or PIR1-PR: 0636, the production strain contains a nucleotide sequence for coding toxin protein and the antibiotic-free plasmid, and the antibiotic-free plasmid contains a nucleotide sequence for coding antitoxin protein; and preferably, the replicon DNA element of the nonreactive plasmid is R6K-gamma. The toxin protein gene of the production strain disclosed by the invention can be stably passaged, has lethality after being induced and can be used for plasmid screening; according to the invention, the positive rate of transforming the nonreactive plasmid into the PIR1-WN:: 0636 strain is more than 80%, and stable production of the plasmid with a high superhelix ratio can be realized.

The invention provides an antibiotic-free host-plasmid TA maintenance system and application thereof, and relates to the technical field of gene engineering. According to the invention, a toxin gene CcdB is accurately integrated to a host escherichia coli chromosome fhuA site through a gene editing technology, and accurate control of toxin expression is realized by adopting an inducible promoter. The antitoxin gene is simplified to a plasmid skeleton, and the toxin inhibition function is maintained through constitutive expression. According to the method, plasmid-free cells are eliminated through arabinose-induced toxin expression, so that stable maintenance of plasmids is realized (the loss rate is lt; and the method completely avoids the use of antibiotics, meets the biological safety requirements, is suitable for large-scale production of vaccines, recombinant proteins and other biological products, and has the advantages of high plasmid yield, low environmental leakage risk, process compatibility and the like.

The invention belongs to the technical field of bioengineering, and relates to a construction method and application of recombinant yarrowia lipolytica for synthesizing plant antitoxin Kauralexin A1. The method is realized by constructing recombinant yarrowia lipolytica. The construction method of the recombinant yarrowia lipolytica for synthesizing the Kauralexin A1 comprises the following steps: overexpressing a codon optimized antitoxin Kauralexin A1 biosynthetic pathway in yarrowia lipolytica, screening adaptive P450 reductase to improve the electron transfer efficiency in a P450 system, and constructing a metabolic pathway scaffold-free multi-enzyme complex to enhance cascade biological catalysis and metabolic flux, so as to obtain the recombinant yarrowia lipolytica for synthesizing the Kauralexin A1. The expression of the mevalonic acid pathway is enhanced, so that a sufficient GGPP precursor is provided for the biosynthesis of the Kauralexin A1. The construction method of the yarrowia lipolytica for biosynthesizing the antitoxin Kauralexin A1, provided by the invention, is simple to operate, and the constructed yarrowia lipolytica can be used for efficiently producing the Kauralexin A1 and has relatively high production and application values.

The present disclosure provides compositions comprising sequence specific endoribonuclease and methods of their use in RNA analysis, RNA synthesis and fingerprinting of RNA molecules. In particular the present disclosure relates to compositions and samples comprising isolated endoribonuclease of Type III toxin-antitoxin systems preferably endoribonucleases of subfamily CptN and subfamily TenpN.

Provided herein are multi-strain population control systems, methods, kits, and compositions. Also provided are methods, systems, kits, and compositions for culturing bacterial cells in multi-strain ecosystems, and temporally arranged multi-strain ecosystems or cultures using a synchronized lysis circuit in combination with multiple toxin / antitoxin systems to cycle continuously over a long period of time.

The application belongs to the field of genetic engineering, and specifically discloses a new sterilization method for realizing precise and efficient sterilization of drug-resistant bacteria by combining CRISPR and a toxin-antitoxin element coupled therewith. The application innovatively uses the guardian RNA element CreTA of CRISPR to improve the stability of the CRISPR-Cas element in the process of sterilizing drug-resistant bacteria, realizes the double sterilization effect of CRISPR and TA, and thus effectively improves the efficiency of the CRISPR sterilization technology in practical application.

The invention discloses a stable and antibiotic-free plasmid expression system and application thereof in preparation of antitumor drugs. The plasmid expression system comprises a replicon, a regulatable promoter, a ribosome binding site RBS and an axe / txe toxin-antitoxin stabilization module, and the plasmid expression system can realize long-term stable expression of exogenous genes under the antibiotic-free condition. The replicon is selected from one or a combination of more of pUC, pMB1, p15A or pSC101; the promoter is selected from one or a combination of more of J23100, J23150 or J23112, and the RBS is selected from one or a combination of more of B0032, B0033 or B0034. According to the invention, an antibiotic-free and lasting plasmid platform is established for salmonella-based therapy, and a framework, synthetic biology and cancer treatment opportunity are provided for reliable gene delivery in vivo.

The utility model discloses a quantitative device for filling antitoxin serum products, which comprises a quantitative liquid guide device and a shell, and the quantitative liquid guide device is arranged in the shell; the quantitative liquid guide device comprises a screw rod, a positioning plate, a sliding column, a positioning frame, an adjusting screw rod, a first adjusting nut, a pressure adjusting spring, a sealing pressing plate, a sealing gasket, a lower sealing ring, a sealing plate and a quantitative liquid guide device body, and the positioning plate is connected between the screw rod and the sliding column; one end of the sliding column sequentially penetrates through the quantitative liquid guide device, the sealing plate and the sealing pressing plate to be connected with the adjusting screw rod, the sealing gasket is connected to the sealing pressing plate and matched with the sealing plate for sealing, the sealing plate is sleeved with the lower sealing ring, and a plurality of transition holes are formed in the sealing plate; the quantifying device for filling the antitoxin serum products is simple in structure, convenient to use and practical, and the amount of antitoxin serum fed every time is controlled by rotating the adjusting nut to move the positioning plate to ascend and descend, so that the use of a metering system is reduced.

The utility model discloses a sealing and cleaning device for packaging antitoxin serum products, which comprises a pulley and a cleaning device, and the cleaning device is arranged on the pulley. The cleaning device comprises a cleaning tank, a discharging pipe, a separating device, a feeding port and a flushing head, the separating device and the feeding port are arranged at the top of the cleaning tank, the discharging pipe is arranged in the cleaning tank, one end of the discharging pipe is matched with the separating device, and the other end of the discharging pipe is connected to the inner wall of the cleaning tank; the plurality of flushing heads are arranged on the side wall of the cleaning tank; the sealing and cleaning device for packaging the antitoxin serum products is simple in structure, convenient to use and practical, cleaned penicillin bottle plugs are rapidly guided out through the separating device, meanwhile, the guided-out penicillin bottle plugs are dewatered through centrifugal force, and the purpose of rapidly guiding out the penicillin bottle plugs after sealing and cleaning is achieved.

This invention relates to an antitoxin protein binding molecule and a kit thereof. The antitoxin protein binding molecule comprises an antibody or an antigen-binding fragment targeting the antitoxin protein. The binding molecule includes a light chain variable region and a heavy chain variable region. The light chain variable region includes a complementarity-determining region (LCDR), which comprises LCDR1 with the sequence SEQ ID NO:1, LCDR2 with the sequence SEQ ID NO:2, and LCDR3 with the sequence SEQ ID NO:3. The heavy chain variable region includes a complementarity-determining region (HCDR), which comprises HCDR1 with the sequence SEQ ID NO:4, HCDR2 with the sequence SEQ ID NO:5, and HCDR3 with the sequence SEQ ID NO:6.

There is provided an antibiotic-free microplasmid comprising a nucleotide sequence encoding an antitoxin protein and a replicon. The skeleton of the non-antibiotic microplasmid is small and is controlled within 1000bp, redundant useless fragments are reduced, a resistance expression cassette is not contained, the utilization rate of a target sequence in the plasmid is improved, and the production burden is reduced. The method has higher efficiency and safety in plasmid DNA-mediated non-viral vector delivery, gene therapy, vaccine immunization, virus production, antibody production and other genetic engineering aspects. The invention also provides a system for producing plasmids based on a toxin-antitoxin system, the bacterial host cell contains a gene expression cassette capable of inducing expression of toxin protein, and the non-antibiotic microplasmids contain a gene expression cassette for expressing antitoxin protein and are used for efficiently maintaining replication and amplification of the non-antibiotic microplasmids.

The invention provides a resistance-free selection marker plasmid, a preparation method, recombinant bacteria and application, and belongs to the technical field of gene engineering. The plasmid comprises an EM7 promoter, a ccdA41 antitoxin gene and an R6K gamma replication origin, the length of the plasmid molecule is less than 600bp, and the plasmid molecule does not contain antibiotic resistance genes. The plasmid is small in molecular weight and free of antibiotic resistance genes, effectively avoids safety risks caused by use of antibiotics, strictly depends on designated host replication of chromosome integration ccdB, meets biological safety requirements and is suitable for safe production of biological products such as vaccines and recombinant proteins. According to the invention, the recombinant bacterium is used for producing the plasmid without the resistance selection marker simply, conveniently and quickly, the success rate is high, and the recombinant bacterium utilizes the GT115-ccdb strain and the plasmid containing the ccdA41 antitoxin gene to realize stable maintenance and expression of the plasmid in a host through a toxin-antitoxin composite regulation mechanism.

The present invention relates to a V. cholerae strain in which a platform has been built to detect integron cassettes. The platform includes a blind reporter system built by inserting an attl integration site into the gene of the toxin of a toxin / antitoxin system, which is inserted in the chromosome of the bacterium; the gene of the corresponding antitoxin and the gene of the intl integrase, both included in plasmids. The invention also relates to V. cholerae strains in which the superintegron (ASI) has been deleted, to the method for obtaining said strains, and to V. cholerae ASI strains in which the platform for capturing integron cassettes has been included. The invention includes methods and kits for detecting integron cassettes by using the strains and platforms built.

The invention discloses a recombinant genetically engineered bacterium for stably producing beta-alanine without antibiotics and inducers and application of the recombinant genetically engineered bacterium, the recombinant genetically engineered bacterium takes Escherichia coli probiotics E15 as a starting strain, and one or more of the following gene editions are carried out: (1) weakening an icd gene; (2) introducing a toxin-antitoxin system hok / sok; (3) knocking out the lacI gene; and (4) overexpression of a temperature-sensitive regulation and control system: the temperature-sensitive regulation and control system is obtained by replacing a lacI gene promoter with a PR promoter and introducing a CI857 repressor protein coding gene expressed by a PL promoter. Finally, the strain constructed by the invention can realize stable and efficient production of beta-alanine in the fermentation process without adding antibiotics and inducers.

The invention relates to the technical field of biology, and discloses a genetically engineered bacterium capable of stably producing 1, 4-butanediol at high yield and application of the genetically engineered bacterium. The genetically engineered bacterium which carries exogenous plasmids and has the capacity of stably accumulating the 1, 4-BDO at high yield outside cells is obtained through metabolic transformation, and compared with an initial strain, the genetically engineered bacterium can better utilize glucose to produce the 1, 4-BDO. The yield of 1, 4-BDO can be increased from 2.45 g / L to 7.59 g / L through the steps of knockout of a dapD gene, a metA gene, a ycA gene, a teB gene, a dcuB gene, a dcuC gene and an amn gene, and introduction of a dapH-patA-dapL gene cluster, a metA-yjcI gene cluster and an fxpK gene. And by introducing a control element TREadhE and a toxin-antitoxin system hok / sok, the fermentation yield of the genetically engineered bacterium can be stably kept at 7.60 g / L under the condition of not adding antibiotics and inducers.

The present invention relates to a V. cholerae strain in which a platform has been built to detect integron cassettes. The platform includes a blind reporter system built by inserting an attl integration site into the gene of the toxin of a toxin / antitoxin system, which is inserted in the chromosome of the bacterium; the gene of the corresponding antitoxin and the gene of the intl integrase, both included in plasmids. The invention also relates to V. cholerae strains in which the superintegron (ASI) has been deleted, to the method for obtaining said strains, and to V. cholerae ASI strains in which the platform for capturing integron cassettes has been included. The invention includes methods and kits for detecting integron cassettes by using the strains and platforms built.

The invention relates to a protein interaction screening system based on a bacterial two-hybrid technology, which comprises a bait protein and prey protein co-expression plasmid pCDFduet1-lambda cI-RNAP, the sequence of which is as shown in SEQ ID No. 1; a reporter plasmid is pACYC184-cIamp; the sequence of the lac promoter-GFP is as shown in SEQ ID No. 2, and the lac promoter-GFP is as shown in SEQ ID No. 2; the positive control plasmid is a pCDFduet1-lambda cI-LG2-RNAP-Gal11p plasmid, and the sequence of the positive control plasmid is as shown in SEQ ID No. 3; the invention further relates to a high-throughput protein interaction screening method of mycobacterium tuberculosis toxin-antitoxin and application of the protein interaction screening system. The protein interaction screening system can realize high-throughput, high-sensitivity and semi-quantitative many-to-many protein interaction analysis by utilizing a flow cytometry sorting technology and three-generation sequencing analysis, effectively solves the problem of false negative caused by non-uniform conversion efficiency of a traditional double-plasmid system, remarkably improves the practicability of high-throughput screening, and has a good application prospect. The large-scale screening bottleneck is broken through, and wide application prospects are achieved.

The invention discloses a linalool production engineering bacterium BYC-JZ-009-linalool-X1 and application of the linalool production engineering bacterium BYC-JZ-009-linalool-X1, and the linalool production engineering bacterium BYC-JZ-009-linalool-X1 is escherichia coli and contains a plasmid pJBEI-MVA-hok / sok-3 and a plasmid pET-linalool casete. According to the invention, by constructing engineering bacteria and optimizing a linalool synthesis route, the yield is obviously increased and exceeds the level of the prior art; a toxin-antitoxin system is introduced, so that the plasmid stability is enhanced, the use of antibiotics is reduced, and the production cost and the environmental burden are reduced; optimized yeast extract powder and peptone are screened, the fermentation formula is improved, and the production efficiency is improved; the optimized fed-batch and extraction process ensures high yield and stability, and is suitable for industrial application; according to the method, dependence on plant resources is reduced, economic benefits and environmental friendliness are achieved, and a feasible scheme is provided for green production of linalool.

The invention discloses a cis-element library for thermophilic microorganism gene expression regulation and application. The cis-element library comprises a constitutive promoter, an inducible promoter and a ribosome binding site (RBS). The invention also discloses application of the element library in gene expression of thermophilic microorganisms. The toxin-antitoxin gene is used as a new selective marker to be applied to genetic manipulation of thermophilic microorganisms by rationally controlling the strength of the promoter. Promoters of different types and strengths are used for replacing natural promoters of genes related to hydrogen production pathways, and hydrogen metabolic pathways in thermophilic microorganisms are modified, so that the yield of hydrogen is improved. And a cis-element library for accurately regulating and controlling gene expression is provided for carrying out basic research on metabolic engineering and molecular biology in thermophilic microorganisms.

The invention discloses a novel gene expression plasmid pTAEP2 capable of realizing efficient and stable overexpression in brevibacillus laterosporus (Brevibacillus laterosporus), and a preparation method of the novel gene expression plasmid pTAEP2 capable of realizing efficient and stable overexpression in brevibacillus laterosporus. The plasmid contains a toxin-antitoxin system gene cassette, so that the plasmid can stably exist in a specific non-mode host; the P2 promoter can drive high-level expression of a target gene; the RepF replicon is contained, so that the plasmid can be effectively replicated in a non-mode host; the composition contains ampicillin antibiotics and erythromycin which are used as selection markers. The invention also provides a method for target protein expression by transforming specific non-mode brevibacillus laterosporus with the pTAEP2 plasmid, the plasmid contains a TraJ element, so that the plasmid is efficiently introduced into a non-mode microbial host including brevibacillus laterosporus in a conjugational transfer manner, the gene introduction process is greatly simplified, and the production cost is reduced. And stable and high expression of a target gene is realized. The invention provides a powerful molecular biological tool for developing efficient and stable gene overexpression research and industrial application, and is beneficial to efficient gene manipulation in the non-model bacteria.

The present invention relates to halophilic microorganisms. In particular, halophilic microorganisms that expresses citramalate synthase. A plasmid encoding an antitoxin-toxin system, and a halophilic microorganism comprising said plasmid are also described, and methods of producing halophilic organisms are also described herein.

The utility model discloses an electricity detection device for producing antitoxin serum products, which comprises a plastic insulating plate, a motor, a conveying screw rod, a limiting baffle plate, a turnover mechanism, an electricity detection mechanism, a mounting bearing and a fixing frame, the plastic insulating plate is mounted on the fixing frame, the motor is mounted on the plastic insulating plate, a power rod is connected with the conveying screw rod, and the limiting baffle plate is mounted on the mounting bearing. One end of the conveying screw rod is connected with the turnover mechanism, the turnover mechanism is connected to the plastic insulating plate through a mounting bearing, the limiting baffle is arranged on one side of the conveying screw rod, and the electric detection mechanisms are arranged on the plastic insulating plate and matched with the conveying screw rod; the electric detection device for producing the antitoxin serum products is simple in structure, convenient and practical, leakage products can be quickly screened out through the electric detection mechanism, meanwhile, the leakage products are sorted out, the purpose of screening while detecting is achieved, and the detection and screening effect is improved.

The invention relates to the technical field of pest control, and particularly discloses a pest control method based on transgenosis, two pairs of toxin / antitoxin genes are crossed and combined in a transgenosis mode and then are respectively inserted into the same site of a pair of homologous chromosomes (two chromosomes) of a pest, and the obtained two homologous chromosomes cannot be separated; if the two homologous chromosomes are separated, the polypide is dead. Male and female transgenic pests comate, half offspring survives, but the offspring dies when the transgenic pests comate with wild pests, so that the purpose of limiting breeding of the pests or replacing wild populations is achieved. The pest control method provided by the invention does not depend on compounds such as tetracycline and the like, is environment-friendly, and improves the fatality rate of offspring of pests, limits the reproduction of the pests, or reduces the carrying and propagation of pathogenic microorganisms and reduces direct or indirect harm caused by the pests in a manner of releasing transgenic pests.

The present disclosure provides compositions comprising sequence specific endoribonuclease and methods of their use in RNA analysis, RNA synthesis and fingerprinting of RNA molecules. In particular the present disclosure relates to compositions and samples comprising isolated endoribonuclease of Type III toxin- antitoxin systems preferably endoribonucleases of subfamily CptN and subfamily TenpN.

Provided is an antibiotic resistance gene-free plasmid-containing production strain. The production strain is a gene-edited strain of the PIR strain, designated as PIR1-WN::0636 or PIR1-PR::0636. The production strain contains a nucleotide sequence encoding a toxin protein and an antibiotic resistance gene-free plasmid, wherein the antibiotic resistance-free plasmid contains a nucleotide sequence encoding an antitoxin protein; preferably, the replicon DNA element for the antibiotic resistance gene-free plasmid is R6K-γ. The toxin protein gene of the provided production strain can be stably maintained during strain passage, and has lethality upon induction, which can be used for plasmid screening. When the provided antibiotic resistance gene-free plasmid is transformed into the PIR1-WN::0636 strain, the positive rate reaches 80% or higher. Moreover, the antibiotic resistance gene-free plasmid can enable the stable production of plasmids with a high supercoiled proportion.