71 results about "Vector (molecular biology)" patented technology

The invention belongs to the technical field of molecular biology, and particularly relates to a CeNF-YA3 gene, an expression vector and application of the CeNF-YA3 gene in vegetable fat regulation and control. The nucleotide sequence of the CeNF-YA3 gene is as shown in SEQ ID NO.1. After the CeNF-YA3 gene is over-expressed, the grease content of arabidopsis seeds and leaves can be remarkably increased; in addition, the invention also provides a series of vectors containing the CeNF-YA3 gene for subcellular localization, yeast hybridization and plant overexpression for the first time, and proves that the CeNF-YA3 protein has a transcriptional activation function, is localized in a cell nucleus and accords with the basic characteristics of transcription factors for the first time. Therefore, the technical scheme of the invention provides scientific guidance for the application of the CeNF-YA3 gene and the protein thereof.

The invention discloses a stropharia rugoso-annulata SRU6 gene promoter, a silent vector containing the promoter and application of silent stropharia rugoso-annulata endogenous genes, and relates to the field of molecular biology and gene engineering. According to the principle of shRNA mediated gene silencing, a silencing vector suitable for gene silencing of stropharia rugoso-annulata is constructed, and meanwhile, a stropharia rugoso-annulata efficient genetic transformation system is established; by utilizing the silent vector and the transformation system established by the invention, the expression quantity of the target gene can be reduced by about 80%, the purpose of gene function research is achieved, and the silent vector and the transformation system have very strong theoretical research and application values.

This application relates to the field of molecular biology, and more specifically to isolated transposases and their use. More specifically to nucleic acids and nucleic acid constructs encoding transposases, nucleic acid sets and nucleic acid set constructs, and compositions, recombinant vectors, recombinant host cells and kits comprising transposases. More specifically to methods for introducing exogenous nucleic acid fragments into the genome of host cells, methods for editing the genome of host cells, and methods for obtaining host cells containing exogenous nucleic acid fragments in their genomes. More specifically to the use of transposases, nucleic acids and nucleic acid constructs, nucleic acid sets and nucleic acid set constructs, compositions, recombinant vectors, or recombinant host cells for introducing exogenous nucleic acid fragment genes into the genome of host cells, or for preparing drugs or formulations for gene therapy, cell therapy, genome research, or stem cell induction and post-induction differentiation.

The invention relates to a Sebastes schlegeli endogenous retrovirus envelope protein Penv, a lentiviral vector and application thereof, and belongs to the field of genetic breeding of molecular biology, and the amino acid sequence of the envelope protein Penv is as shown in SEQ ID NO.1. The invention further provides an in-vitro assembly system, a transformant and a kit containing membrane fusion protein particles and lentiviral particles of the envelope protein Penv and application of the envelope protein Penv. VSVG protein is replaced with Penv protein from sclerobone fish, so that the transduction efficiency of the lentiviral particles to sclerobone fish cells is effectively improved, and the transduction efficiency of the lentiviral particles to the sclerobone fish cells is improved. And efficient gene delivery is realized. Meanwhile, as the endogenous Env protein, the Env protein overcomes the immunological rejection of a host to the exogenous Env protein, and also has the potential of in-vivo application.

The present application relates to the field of molecular biology, in particular to a recombinant human heparin N-deacetylase encoding gene, a recombinant vector, a recombinant strain and application thereof, the recombinant human heparin N-deacetylase constructed in the expression process shows excellent stability and activity, has the ability of specific catalysis of deacetylation of acetyl group in heparin precursor, and can effectively generate deacetylated heparin precursor. This property significantly improves the conversion efficiency and reduces the generation of by-products. The present application optimizes the codon of the recombinant human heparin N-deacetylase, and uses the promoter P AOX1 or P GAP to regulate the expression, so that the expression level of the recombinant human heparin N-deacetylase in Pichia pastoris is significantly improved, and the obtained GS115-AOX1-hNDase and GS115-PGAP-hNDase strains successfully realize the secretory expression of the target gene, providing a new strategy and method for the biosynthesis of heparin.

The present invention belongs to the field of molecular biology and genetics, and in particular relates to a method for editing a bovine gene based on Pro-iCHI. The present invention provides a method for editing a bovine gene based on Pro-iCHI. Protamine is transiently expressed in gene-edited b-haSCs, which are then injected to mature oocytes to obtain reconstructed embryos. Protamine can eliminate abnormal DNA methylation resulting from oocyte intracytoplasmic haSCs injection and enable the nucli to compress into sperm-like structures, and the obtained bovine Pro-iCHI embryos can successfully develop into blastocysts, with a blastocyst rate comparable to that of the embryos obtained by in vitro fertilization. Moreover, in the present invention, a protamine-encoding gene is inserted into a Saccharomyces cerevisiae protein expression vector for transient expression, which ensures that abnormal DNA methylation is erased, without integration into the genome resulting in the insertion of exogenous genes.

This invention discloses a novel site-specific recombinase-mediated multisite chromosome editing tool and its applications. The invention provides a complete product suite for integrating or continuously integrating the genome of a host cell, comprising: a circular vector 1 composed of the following elements: loxF1, nox, ori, resistance gene, lox71, P3, P1, P2; and / or a circular vector 2 composed of the following elements: lox66, nox, ori, resistance gene, loxF2, P3, P1, P2; and a circular vector 3 for expressing Nigri and Cre recombinases; and a site-specific knock-in tool for knocking in the lox66-nox fragment at the desired integration site in the host cell genome; P1 and P2 are adapters, used to insert the DNA fragment to be integrated, and P3 is a universal primer. This invention enables rapid and continuous genome integration using simple molecular biology operations.

The present invention relates to the technical field of molecular biology, and in particular, to an anti-CD40 nanobody, and a preparation method therefor and use thereof. The anti-CD40 nanobody provided by the present invention has three unique complementarity determining regions, namely CDR1, CDR2, and CDR3. The present invention further provides a sequence encoding the nanobody or its VHH chain, a corresponding expression vector, and a host cell capable of expressing the nanobody. The present invention includes immunizing an alpaca by constructing a CD40 antigen to obtain a PBMC of the alpaca and constructing a vector; screening an anti-CD40 antibody through phage display; and finally obtaining a highly sensitive and specifically binding antibody through functional assays and sequencing result analysis. The nanobody provided by the present invention can specifically recognize and bind to CD40.

The invention discloses a ribosome group or selective ribosome group detection and analysis method based on an immune principle, and relates to the field of molecular biology and genomics. Comprising the following steps: sample preparation and cross-linking: treating cells by using a translation inhibitor to suspend ribosome translation, and then cross-linking RNA-protein in the cells by using a membrane permeable cross-linking agent; then cracking the cells, digesting RNA of a cracking product by using ribonuclease, and retaining ribosome and mRNA fragments protected by the ribosome; immune enrichment: carrying out specific immune enrichment on the digested sample by using an antibody combined with a carrier, and eluting to obtain a ribosome-mRNA compound; then decrosslinking and removing protein, extracting mRNA fragments and carrying out phosphorylation treatment so as to add joints, build a library and carry out sequencing; and finally, library construction, sequencing and bioinformatics analysis are carried out. The method has the advantages of high-specificity enrichment, accurate positioning of translation sites, wide application range and accurate sequence information.

This application relates to the fields of gene therapy and molecular biology. More specifically, the present invention relates to an isolated altered VP1 protein of adeno-associated virus serotype 5 (AAV5) capsid comprising one or more amino acid substitutions as compared to the VP1 protein of wild-type AAV5 capsid, which increase transduction efficiency, as well as to a capsid and a vector based thereon.

The invention relates to a construction method of an OriCiro cell-free cloning system based on an RCR technology, and belongs to the technical field of molecular biology. The construction method comprises the following steps: expression and purification of high-activity Tn5 transposase, transposon-mediated oriC insertion, RCR amplification, traceless excision of oriC transposon and repair cyclization. The construction method provided by the invention provides a rapid, reliable and mutation-burden-free new genome construction and rescue path for the existing research, can solve the difficult problem of construction of large-size and toxic DNA sequence vectors for gene therapy, shortens the research and development and production periods, and improves the production efficiency. Therefore, a powerful tool is provided for the fields of preparation of mRNA vaccines or therapy plasmids, construction of gene therapy vectors, supply of pharmaceutical-grade plasmid DNA, research of a virus molecular mechanism and the like.

The invention relates to the technical field of molecular biology, in particular to an antibacterial peptide SAV-546 as well as a construction method and application thereof, and the construction method of the antibacterial peptide SAV-546 comprises the following steps: obtaining a target gene segment through gene cloning; constructing an expression recombinant vector by using the target gene segment to obtain a positive recombinant plasmid; and carrying out prokaryotic induced expression on the positive recombinant plasmid to obtain the antibacterial peptide SAV-546. The antibacterial peptide SAV-546 disclosed by the embodiment of the invention is not easy to induce bacteria to generate drug resistance, the construction method of the antibacterial peptide SAV-546 is simple, convenient and efficient, the cost is low, the antibacterial peptide SAV-546 can be widely applied to preparation of antibacterial drugs, medical dressings and food preservatives, and a new scheme is provided for solving the problem of clinical drug-resistant bacterial infection.

The invention relates to the technical field of molecular biology, in particular to a construction method of a J774A.1 cell line with a baz2b gene knocked out. The invention provides a construction method of a J774A.1 cell line with a baz2b gene knocked out. The construction method comprises the following steps: constructing a J774A.1 cell line for expressing Cas9 protein; the recombinant vector contains the sgRNA; a recombinant vector, pMD2. G and psPAX2 are co-transfected into cells, and then the cells, a J774A.1 cell line for expressing Cas9 protein and puromycin antibiotic are screened to obtain a J774A.1-KO baz2b cell line with inactivated baz2b function, and the J774A.1 cell line with the baz2b gene knocked out is obtained. The cell line can regulate the formation of a macrophage extracellular trapping net.

The invention relates to the field of genetic engineering and molecular biology, in particular to a nucleic acid construct, and a vector, a host cell, a viroid particle, a vaccine and a pharmaceutical composition containing the nucleic acid construct. Furthermore, the invention also provides application of the nucleic acid construct vector, the host cell, the viroid particle, the vaccine and the pharmaceutical composition in prevention and / or treatment of virus (e.g., SARS-CoV-2) infection or diseases and / or symptoms caused by virus infection.

The invention relates to the fields of genetic engineering and molecular biology, in particular to a method for optimizing a nucleotide sequence of an expression protein in a host cell and a method for optimizing a nucleotide sequence of an expression protein in a Vero cell. The invention also relates to nucleic acid molecules, vectors, host cells and viroid particles comprising the optimized nucleotide sequences obtained by said method.

More specifically, relating to the field of molecular biology, this invention relates to isolated nucleases and their uses. More specifically, to nucleases, guide RNAs and nucleic acid constructs encoding the nucleic acid constructs thereof, as well as compositions containing nucleases, recombinant vectors, recombinant host cells and kits. More specifically, to methods for introducing double-strand breaks into targeted genes in host cells, methods for deleting, replacing or inserting targeted genes in host cells, and methods for obtaining host cells into which targeted genes are deleted, replaced or inserted. More specifically, to the use of nucleases, nucleic acids and nucleic acid constructs encoding nucleases, guide RNAs and their nucleic acid constructs, compositions, recombinant vectors, or recombinant host cells for introducing double-strand breaks into targeted genes in host cells, deleting, replacing or inserting targeted genes in host cells, and preparing drugs or preparations.

The application discloses a plant-expressed varicella-zoster virus gB recombinant antigen and a preparation method, product and application thereof, and belongs to the technical field of plant molecular biology and vaccine engineering. The technical problems to be solved are that the existing VZV gB protein has low heterologous expression, poor stability and insufficient antigen activity; the existing VZV vaccine has high production cost, is difficult to popularize and has a single antigen with an immune escape risk; and the plant expression system is not mature in VZV gB antigen production, and lacks a complete preparation scheme and related products. The technical solution is that the natural VZV gB protein is modified, a furin protease cleavage site is knocked out, and a H527P mutation is introduced, related nucleic acid molecules, vectors and plant source cells are constructed, high-activity antigens are obtained through culture and purification, and the high-activity antigens are applied to the preparation of VZV drugs / vaccines, and have the advantages of low cost, high safety and strong immunogenicity.

The invention discloses a bovine SLC27A6 gene promoter vector as well as a construction method and application thereof, and belongs to the field of molecular biology and genetic engineering. According to the invention, the bovine SLC27A6 gene promoter SLC27A6-A-SLC27A6-E of which the nucleotide sequences are respectively shown as SEQ ID NO.1-SEQ ID NO.5 is obtained through cloning, and bovine SLC27A6 gene promoter vectors are respectively constructed by taking a pGL4.10 vector as a skeleton vector. Experimental results show that in MAC-T cells, the bovine SLC27A6 gene promoter vector constructed by the invention can exert promoter activity and regulate and control downstream gene expression. The invention provides a new biological material for regulation of gene expression in bovine cells and genetic breeding of the targeted SLC27A6 gene, and has a wide application prospect.

The invention discloses a DNA structure for saccharomyces cerevisiae single-cell barcode labeling and chromatin state conjoint analysis and application thereof, and belongs to the technical field of molecular biology and single-cell multi-omics analysis. From the 5'end to the 3 'end, a fluorescent protein gene, a Tn5ME structure, a reverse transcription primer binding region, a CYC1 terminator fragment and a linearized shuttle plasmid skeleton are subjected to homologous recombination, and a pCM carrier is obtained; after NdeI / BamHI enzyme digestion, oligonucleotides with random sequences, which are subjected to annealing, amplification, Klenow filling-in and enzyme digestion, are connected, and transformation extraction is carried out, so that the DNA structure for saccharomyces cerevisiae single-cell barcode labeling and chromatin state combined analysis is obtained. On the basis of the constructed DNA structure, ATAC-seq and a bar code reverse transcription method are combined, important data related to the transcription state and chromatin state of a single cell can be obtained at the same time through bar code information, single cell pedigree tracking of the saccharomyces cerevisiae is achieved, and research on single cell epigenomics of the saccharomyces cerevisiae and other microorganisms is promoted.

The invention discloses a method for improving canker resistance of citrus by using a CsFKBP16-4 gene, and relates to the field of molecular biology, the transcriptional level of the FKBP16-4 gene in citrus plants is reduced by interfering RNA, and the nucleotide sequence of the interfering RNA is as shown in SEQ ID NO: 1. A part of coding sequence of the CsFKBP16-4 gene is used as an interference sequence, is introduced into an interference vector, and is integrated into a citrus genome through an agrobacterium-mediated genetic transformation method, so that the canker resistance of citrus is effectively improved. The canker attack degree of the obtained transgenic plant can be reduced to 28.3% of that of the existing citrus, the canker symptom of the transgenic citrus is remarkably relieved, and the canker resistance of the citrus is improved. In addition, transgenic citrus plants can directly inhibit proliferation of pathogenic bacteria causing canker, the canker can be ensured to be inhibited essentially, another way for citrus canker-resistant breeding is opened up, and development of citrus canker-resistant breeding is promoted.

The application provides a broad-spectrum monoclonal antibody of enterovirus A population and application thereof, and belongs to the technical field of molecular biology and immune analysis. The monoclonal antibody comprises the following six CDR regions: the amino acid sequences of the CDR regions of the heavy chain are respectively shown in SEQ ID No. 1-SEQ ID No. 3; and the amino acid sequences of the CDR regions of the light chain are respectively shown in SEQ ID No. 4-SEQ ID No. 6. The application also provides a nucleotide fragment encoding the monoclonal antibody, a recombinant vector and an engineering cell line comprising the nucleotide fragment, an antibody conjugate and their uses. The monoclonal antibody is a non-neutralizing antibody of IgG1 subtype, can be combined with enterovirus A population viruses in a broad spectrum, and can be used for preparing a reagent or / and a kit for detecting enterovirus A population viruses, and for preparing a medicine for inhibiting, preventing or / and treating enterovirus A population viruses.

The application provides a preparation method and application of an RNA cell-free synthesis preparation, and belongs to the technical field of molecular biology. The preparation method of the RNA cell-free synthesis preparation is mainly as follows: taking Escherichia coli as a chassis strain, using a pET-28a vector to construct a recombinant strain for expressing T7 RNA polymerase, the T7 RNA polymerase has enzymatic activity at 50-60 DEG C; the bacterial cells are collected by induction culture and are subjected to ultrasonic lysis, and the lysis solution is treated to obtain the preparation for RNA cell-free synthesis. The preparation provided by the application naturally contains various cofactors, and the RNA polymerase does not need to be extracted and purified, so that the production cost is reduced.

This application relates to the field of molecular biology, and more specifically to isolated transposases and their use. More specifically to nucleic acids and nucleic acid constructs encoding transposases, nucleic acid sets and nucleic acid set constructs, and compositions, recombinant vectors, recombinant host cells and kits comprising transposases. More specifically to methods for introducing exogenous nucleic acid fragments into the genome of host cells, methods for editing the genome of host cells, and methods for obtaining host cells containing exogenous nucleic acid fragments in their genomes. More specifically to the use of transposases, nucleic acids and nucleic acid constructs, nucleic acid sets and nucleic acid set constructs, compositions, recombinant vectors, or recombinant host cells for introducing exogenous nucleic acid fragment genes into the genome of host cells, or for preparing drugs or formulations for gene therapy, cell therapy, genome research, or stem cell induction and post-induction differentiation.

The present application relates to a kind of Sebastes schlegelii endogenous retrovirus envelope protein P_env, lentivirus vector and its application, belong to the genetic breeding field of molecular biology, the amino acid sequence of the envelope protein P_env is as shown in SEQ ID NO.1.The present application simultaneously provides the membrane fusion protein plasmid of containing the envelope protein P_env, the in vitro assembly system of lentivirus particle, transformant and kit, and the application of the envelope protein P_env, the P_env protein of the present application is replaced VSVG protein using hard fish source, will effectively improve the transduction efficiency of lentivirus particle to hard fish cell, realize efficient gene delivery.Meanwhile, as endogenous Env protein, overcome the immune rejection of host to exogenous Env protein, also have the potential of in-vivo application.

The present application relates to the fields of gene therapy and molecular biology. More specifically, the present invention relates to an isolated modified capsid protein VP1 from adeno-associated virus serotype 9 (AAV9) comprising one or more amino acid substitutions compared to the wild-type AAV9 capsid protein VP1, which substitutions increase the efficiency of production (assembly) of the vector based on recombinant adeno-associated virus serotype 9 (rAAV9), to a capsid and a vector based on the above VP1, as well as to uses thereof.

The application belongs to the technical field of insect resistance research, and discloses a method for enhancing the insect resistance of poplar by using a leaf senescence related gene NAC109, which is characterized by comprising the following steps: (1) obtaining a CDS sequence of a PtNAC109 gene; (2) designing a gRNA sequence; (3) constructing a targeted PtNAC109 gene editing vector plasmid; (4) preparing a genetic transformation agrobacterium infection solution; (5) using agrobacterium to infect poplar plants; (6) screening resistant callus; (7) inducing, prolonging and rooting the resistant shoots; (8) verifying the gene editing effect by using molecular biology means; and (9) obtaining a new poplar gene editing germplasm with enhanced insect resistance.

The invention belongs to the field of molecular biology and immunology, and particularly relates to an anti-CD3 nano antibody capable of activating the activity of T cells as well as a preparation method and application of the anti-CD3 nano antibody. The invention provides a preparation method of the anti-CD3 nano antibody. The preparation method comprises the following steps: constructing a CD3 plasmid and an overexpression cell line, and immunizing alpaca to obtain peripheral blood mononuclear cells (PBMC) of the alpaca; extracting RNA (Ribonucleic Acid) and carrying out reverse transcription to obtain cDNA; carrying out PCR amplification on the antibody gene segment and cloning the antibody gene segment to a phage display vector; and electrically transforming the vector into SS320 competent cells to obtain the nano antibody library. After two rounds of panning, detecting and screening an antibody clone capable of being combined with the CD3 protein through ELISA (Enzyme-Linked Immunosorbent Assay); an antibody sequence is selected through sequencing, and an antibody protein is expressed by using a mammalian cell high-throughput expression system; and then detecting and screening the antibody which is only specifically combined with the CD3. The method disclosed by the invention is simple, convenient and rapid to operate, and can be used for preparing active antibodies for treating potential tumors and proliferative diseases.

The application belongs to the technical fields of plant molecular biology and genetic engineering, and relates to a poplar PtoNAC43 transcription factor gene and application thereof. The nucleotide sequence of the gene is shown as SEQ ID NO:1, and the gene encodes a transcription factor protein with a NAC domain. A transgenic plant is obtained by constructing an overexpression vector and introducing the vector into a poplar. The results show that overexpression of the gene can reduce plant height, biomass and growth potential, and affect photosynthesis parameters, endogenous hormone levels and tissue structure. The application can be used for plant growth regulation and dwarf plant cultivation.

The application discloses a carp U6 promoter which is cross-species function-conserved and drives long-fragment sequence expression and an application thereof, and belongs to the technical field of genetic engineering and molecular biology. The existing U6 promoter is strong in species specificity, low in activity in non-mammals such as fish, and mainly suitable for short-fragment RNA transcription, and is difficult to drive long-fragment target gene expression, thereby limiting complex gene editing and transgenic aquatic animal creation. The application provides a U6 promoter derived from a carp, and the nucleotide sequence is shown in SEQ ID NO:1. The promoter has cross-species function-conservation, and can drive long-fragment target gene transcription. An expression cassette and a recombinant vector constructed based on the promoter can realize high-efficiency expression of long-fragment target genes in fish cells and mammal cells. The promoter and related biological materials derived therefrom can provide efficient and cross-species applicable key technical tools for fish gene function research and editing breeding.

The invention relates to the technical field of molecular biology, in particular to a carrier for preparing circular RNA, application of the carrier and a method for preparing the circular RNA. The vector comprises a 5'end sequence connected with ribozyme and a 3 'end sequence connected with ribozyme, a target sequence needing to be looped is arranged between the 5' end sequence connected with ribozyme and the 3 'end sequence connected with ribozyme, and accurate looping of in-vitro precursor RNA can be achieved. The vector provided by the invention can be used for preparing circular RNA, is simple in process, and can be used for synthesizing circular RNA vaccines and drugs.