9 results about "Shuttle vector" patented technology

This invention discloses a recombinant adenovirus vaccine targeting the Tp0326 antigen and its preparation method, comprising a recombinant adenovirus vector and the Tp0326 antigen expressed therein. The recombinant adenovirus vector uses a replication-defective human adenovirus type 5 (Ad5) as a backbone, inserting the full-length Tp0326 antigen gene of *Treponema pallidum* into the adenovirus shuttle vector pshuttle-IRES-rGFP-1, and obtaining it through homologous recombination with the adenovirus backbone plasmid pAdEasy-1. This invention, using a replication-defective Ad5 as a vector, provides a recombinant adenovirus vaccine with high safety and efficient expression of the full-length Tp0326 antigen. The expressed antigen has a natural conformation, retains the ECL4 immunodominant epitope, and is more likely to induce a specific immune response, resulting in stronger immunogenicity. The preparation method involves constructing a recombinant adenovirus vector through homologous recombination, packaging it in HEK293T cells, and purifying it by CsCl density gradient centrifugation to obtain a high-purity, high-titer recombinant adenovirus vaccine suitable for large-scale production. The recombinant adenovirus vaccine can be administered via intramuscular or intranasal injection.

The invention discloses a green fluorescence report system for screening a Listeria monocytogenes quorum sensing inhibitor as well as a construction method and application of the green fluorescence report system. The report system is constructed through the following steps: synthesizing a DNA fragment according to a listeria monocytogenes AGR system pII promoter sequence, connecting the DNA fragment with a green fluorescent protein pEGFP plasmid, amplifying an EGFP-pII target fragment through PCR, and connecting the target fragment to a pKSV7 shuttle vector plasmid through homologous recombination to obtain a pKSV7-EGFP-pII plasmid, and then transferring the gene into a competent cell of listeria monocytogenes ATCC BAA-679, and screening to obtain a report system. The report system disclosed by the invention can be used for intuitively evaluating the expression condition of the quorum sensing system of the listeria monocytogenes and screening out the quorum sensing inhibitor.

The present disclosure provides a recombinant adenovirus for expressing African swine fever virus (ASFV) EP153R-EP402R protein and a construction method thereof, and belongs to the technical field of genetic engineering. In the present disclosure, a recombinant adenovirus vector pAD-CMV-EGFP-EP153R-EP402R is obtained through a series of intermediate processes using a recombinant adenovirus shuttle vector pENTRE-EGFP-TOPO; the recombinant adenovirus vector is linearized to transfect AD293 cells, a recombinant virus is screened according to cytopathy formed by adenovirus infection, an adenovirus packaging process is achieved, and the recombinant adenovirus for expressing ASFV EP153R-EP402R protein is obtained, laying a foundation for the construction of a recombinant adenovirus vaccine for expressing the ASFV EP153R-EP402R protein.

The invention belongs to the technical field of information, and discloses a copper inducible plasmid which comprises a yeast shuttle vector, a Candida glabrata-derived CEN / ARS element loaded on the yeast shuttle vector, a copper ion inducible promoter loaded on the yeast shuttle vector and a target gene, the target gene is connected to the downstream of the copper ion induced promoter. After being transfected into candida glabrata, the plasmid can show the following advantages: the plasmid is stable, the stability of host bacteria cannot be influenced on the basis of controllable expression of the dosage of an inducer, and simultaneous expression of various target proteins can be realized on the basis of loads of different target genes of the plasmid. Meanwhile, the invention also discloses an inducible expression method of the target gene.

The application discloses application of a broad-spectrum type III esterase derived from Rhodococcus in phthalate ester pollution bioremediation, and overcomes the limitation of low degradation efficiency of a natural strain; through genetic engineering technology, the esterase 5359 gene is cloned into a Rhodococcus-E.coli shuttle vector pNV18 to construct a recombinant Rhodococcus engineering bacterium WT-pNV18-5359 overexpressing the esterase. Experimental results show that the engineering bacterium significantly improves the degradation performance: in single PAEs degradation test, the degradation rates of long-chain PAEs such as DEHP, DnOP and DiNP are respectively increased by 15.71%, 13.99% and 12.82% compared with the wild type; in a mixed PAEs system, the degradation of long-chain PAEs is obviously superior, and the engineering bacterium has good genetic stability and does not affect growth. Through genetic engineering to strengthen the application of known enzymes, efficient and stable PAEs pollution bioremediation is realized.

The invention discloses an engineered nitrogen-fixing vibrio natrieae and a preparation method and application thereof, and relates to the field of synthetic biology, a nitrogen-fixing gene nif gene cluster is cloned and assembled on an escherichia coli-yeast shuttle vector to form recombinant plasmids, the plasmids are transformed and extracted by escherichia coli and then transformed into wild vibrio natrieae, and the engineered nitrogen-fixing vibrio natrieae is obtained. And obtaining the nitrogen-fixing vibrio natriivibrio. The invention further discloses the obtained engineered nitrogen-fixing vibrio natriticus and application thereof. The invention develops novel vibrio natriticus capable of efficiently fixing nitrogen in a broad spectrum as well as a preparation method and application of the novel vibrio natriticus.

To provide plasmid DNA with a novel replication origin that can stably introduce foreign genes into actinomycetes. [Solution] The plasmid vector according to the present disclosure comprises (a) a nucleotide sequence having a specific sequence, or (b) a nucleotide sequence in which one or more nucleotides are substituted, deleted and / or inserted in the nucleotide sequence having the specific sequence.

This invention provides a chimeric adenovirus vector Ad5F35 and a recombinant CCR5-32 mutant gene, its construction method, and its application, belonging to the field of gene therapy technology. The construction method provided by this invention includes the following steps: (1) inserting the CCR5Δ32 mutant gene into the multiple cloning site of a shuttle vector to obtain a shuttle plasmid; (2) co-transfecting cells with the shuttle plasmid and the Ad5F35 adenovirus backbone plasmid to generate an Ad5F35 chimeric recombinant adenovirus capable of expressing the CCR5Δ32 gene. By inducing the inability of the CCR5 protein on the host cell to be normally expressed on the cell membrane surface, the binding of HIV-1 gp 120 to the CCR5Δ32 mutant gene is effectively prevented, thus preventing the HIV-1 virus from entering the host cell for replication, thereby achieving the goal of treating AIDS.

The invention provides a resistance marker-free auxotrophic bacillus subtilis and a construction method and application thereof, the newly constructed strain takes bacillus subtilis 168 as a starting strain, indole-3-glycerophosphate synthase gene trpC of the bacillus subtilis 168 is knocked out, synthesis of tryptophan in the growth and metabolism process of the strain 168 is eliminated, and the resistance marker-free auxotrophic bacillus subtilis is obtained. The shuttle vector YH46 is constructed by taking pMD-19T as a framework, so that the shuttle vector YH46 can normally grow only in the presence of exogenous tryptophan, and meanwhile, a universal auxotroph shuttle vector YH46 is constructed by taking pMD-19T as the framework, so that convenience is provided for high-efficiency expression of genes in bacillus subtilis; the tryptophan auxotrophic strain BS-TR is constructed through double exchange by using a homologous recombination principle, the construction process is simple and easy to implement, and the strain BS-TR does not contain a resistance gene selection marker, can be used as a food-grade engineering bacterium, and can be used for intracellular expression or extracellular secretion of target protein for production.