175results about How to "High homology" patented technology

The present invention relates generally to synthetic genes for modifying endogenous gene expression in a cell, tissue or organ of a transgenic organism, in particular a transgenic animal or plant. More particularly, the present invention provides novel synthetic genes and genetic constructs which are capable of repressing delaying or otherwise reducing the expression of an endogenous gene or a target gene in an organism when introduced thereto.

Disclosed and claimed is a method for preparing a normalized sub-divided library of amplified cDNA fragments from the coding region of mRNAs contained in a sample. The method includes the steps of: a) subjecting the mRNA population to reverse transcription using at least one cDNA primer, thereby obtaining first strand cDNA fragments, b) synthesizing second strand cDNA complementary to the first strand cDNA fragments by use of the first strand DNA fragments as templates, thereby obtaining double stranded cDNA fragments, c) digesting the double stranded cDNA fragments with at least one restriction endonuclease, the endonuclease leaving protruding sticky ends of similar size at the termini of the DNA after digestion, thereby obtaining cleaved cDNA fragments, d) adding at least two adapter fragments containing known sequences to the cleaved cDNA fragments obtained in step c), the at least two adapter fragments being able to bind specifically to the sticky ends of the double stranded cDNA produced in step c), the one adapter fragment being able to anneal to the primer having formula I in step f), the second adapter fragment being a termination fragment introducing a block against DNA polymerization in the 5'->3' direction setting out from said termination fragment and the termination fragment being unable to anneal to any primer of the at least two primer sets in step f) during the molecular amplification procedure, the at least two adapter fragments being ligated to the cleaved cDNA fragments obtained in step c) so as to obtain ligated cDNA fragments, e) sub-dividing the ligated cDNA fragments obtained in step d) into 4n1 pools where 1<=n1<=4, and f) subjecting each pool of ligated cDNA fragments obtained in step e) to a molecular amplification procedure so as to obtain amplified cDNA fragments, wherein is used, for an adapter fragment used in step d), a set of amplification primers having the general formula Iwherein Com is a sequence complementary to at least the 5'-end of an adapter fragment which is ligated to the 3'-end of a cleaved cDNA fragment, N is A, G, T, or C, the one primer having the general formula I where n1=0, and the second primer having the general formula I where 1<=n1<=4, the second primer being capable of priming amplification of any nucleotide sequence ligated in its 3'-end to the adapter fragment complementary in its 5'-end to Com.

The invention discloses a CRISPR (clustered regularly interspaced short palindromic repeat)/Cas9 recombinant lentiviral vector containing gRNA sequence specifically targeting CCR5 and application thereof. A lentivirus of the CRISPR/Cas9 recombinant lentiviral vector containing gRNA sequence specifically targeting CCR5 gene Delta 32 region is constructed that the lentivirus can introduce cells into a CRISPR/Cas9 system specific to CCR5, double-chain breakage occurs to a specific site of CCR5 gene, a random mutation is introduced to a breakage site after repairing by means of nonhomogeneous recombinant terminal binding, and the mutation rate reaches 90% and above. As gRNA is a nonhomogeneous region of CCR5 and CCR2, detection shows that the missing efficiency of the two gRNAs is lower than 0.2%. Cells modified via the recombinant lentivirus have significantly decreased efficiency of HIV (human immunodeficiency virus) infection. The system is quick to construct, simple and low in price, and is applicable to gene therapy of acquired immune deficiency syndrome.

The present invention relates to α-amylase variants that are stabilized to dimerization and/or multimerization, in particular at elevated temperatures or high pH, by point mutagenesis of positively polarized or charged or neutral surface amino acids to give more negatively polarized or charged amino acids. The invention further relates to methods of increasing the stability of an α-amylase to dimerization and/or multimerization brought about by electrostatic interactions whereby at least one amino acid residue on the surface of the starting molecule, which makes a neutral or positively polar or charged contribution to the electrostatic potential of said molecule, is replaced with a more negatively polar or negatively charged amino acid residue. The α-amylase variants obtained thereby exhibit better stability to influences of the solvent, increased processivity, and are suited for numerous industrial areas of use, in particular as active ingredients in detergents and cleansers.

The invention discloses a broad-spectrum vibrio subunit vaccine for preventing fish pathogenic vibrio infection and a preparation method, belonging to the technical field of biology. The fusion protein OmpK-FlaA of an outer membrane protein OmpK with a conserved structure and a flagellin protein FlaA on the vibrio parahaemolyticus surface serves as an antigen component of the vaccine. The method is characterized by comprising the following steps of: superposing and extending Ppolymerase chain reaction (PCR) to the ompK and flaA genes of the vibrio parahaemolyticus to obtain a fusion protein gene flaA-ompK; constructing flaA-ompK-pET-28a recombinant plasmids; and obtaining the fusion FlaA-OmpK with high purity through exogenous induction expression and purification. The vaccine prepared bythe invention is safe and nontoxic and has no side effects; injection immune can be adopted; and the immune can also be realized by taking enteric microsphere vaccine orally; cross immunity protection can be acted to fish pathogenic vibrio (vibrio parahaemolyticus, vibrio alginolyticus, vibro harveyi, vibrio anguillarum and vibrio vulnificus).

The invention is about the clone, recombination and the salt tolerant analysis of the GhZFP1gene of the zinc finger structure of the CCCH type in the cotton. The invention is to extract the total RNA from the cotton leaves after treating by the 0.3mol/L NaCl to separate the mRNA and construct the cDNA library, then to select the gene encoded the GhZFP1 protein of the zinc finger of the CCCH type. The tobacco transformed by the gene can grow in the concentration of 200mmol/L salt; and the paddy can grow in the in the concentration of 100mmol/L salt. So it can improve the haloduric ability of the plants to plant the plants in the alkaline soil.

The invention discloses a mutant fibroblast growth factor (FGF-21) and use of the mutant fibroblast growth factor in treating endocrine diseases. The amino acid sequence of the fibroblast growth factor-21 disclosed by the invention is shown as SEQ ID NO: 2, and the sequence of a gene for encoding the mutant fibroblast growth factor is shown as SEQ ID NO: 1. According to the invention, a wild typeFGF-21 is used as a template, two arginine (Arg) residues are introduced through a downstream primer, and a mutant is obtained through polymerase chain reaction (PCR). The strong basicity and positive charges in the physiological condition of Arg are mainly utilized so that the isoelectric point of FGF-21 is up-regulated, and FGF-21 binding to the surfaces of cell membranes is facilitated. The results of an animal experiment show that the mutant FGF-21 disclosed by the invention can more effectively reduce the blood glucose level in an animal, and furthermore, the mutant disclosed by the invention has the advantages of fast onset of drug action, lasting drug effect and the like in reducing the blood glucose level. The mutant GF-21 disclosed by the invention can be used as a medicine to treat endocrine diseases such as diabetes, metabolic syndrome, lipid metabolism disorder and the like.

A method useful for stable expression of an exogenous gene, introduced into a plant, is provided. By introducing an exogenous gene and 5' upstream sequence of tobacco dehydogenase concurrently, stable expression of the exogenous gene introduced into a plant can be achieved.

The invention relates to an injectable implant and a preparation method thereof. The implant is amnion microparticles mixed in a sodium chloride phosphate physiological solution with a pH of 7.0-7.5. The preparation method comprises the following steps of disinfection processing, decellularization processing, modification processing, and amnion microparticle preparation, and in particular, comprises: taking amnion microparticles as a component A and a sodium chloride phosphate physiological solution as a component B, performing vibration and well mixing according to a mass volume ratio of 20:1-100:1 to prepare an amnion microparticle suspension with a concentration of 20-100 mg/mL by high-speed microjet equipment, wherein the particle size of the amnion microparticles is 50-200 microns. The injectable implant of the invention maintains the natural three dimensional stereo structure and a lot of natural active components of human amnion, reduces immunological rejection reaction caused by raw material source after clinical application of cosmetic injection products, is suitable for injection into deep subcutaneous dermal layer to subcutaneous superficial layer to repair moderate to severe wrinkles or folds, and has good cosmetic repair effect.

The invention discloses a DNA extraction method of Duck Enteritis Virus genome and the genome sequence. Firstly, the pyrolysis of chicken embryo fibroblast which is infected with DEV is processed by hypertonic buffer; then the nuclease is added for the digestion of cell DNA; and the virus genome DNA is obtained through phenol chloroform extracting method. The obtained sample is broken randomly into 2kb to 2.5kb fragments; the pUC19 vector is linked to transform host E. coli and construct genomic library. The positive clones in the library are randomly sequenced; the total sequence length has 6 times coverage rate for the virus genome. The sequence result is assembled and clustered to obtain the whole DEV genome sequence with the length of156512bp. The analysis shows that the genome structure is the same with that of the members of Varicella virus in Alpha- herpesus subfamily, which provides theoretical basis for the classification of DEV.

The invention belongs to the technical field of genetic engineering, and relates to recombinant hexamer protein A as well as the constitution method and the application thereof. The recombinant hexamer protein A has an amino acid sequence represented by SEQ ID NO.1 in a sequence list. The recombinant hexamer protein A has higher activity than those of the reported recombinant protein A and the natural protein A, and has high yield and relatively simple purification process, thereby preventing the risk of pathogenic bacteria of naturally extracted protein. Additionally, the recombinant hexamer protein A suits large-scale production.

A method of detecting a mutation is provided that uses Tm analysis and is excellent in detection sensitivity. A detection probe consisting of a polynucleotide complementary to a sequence to be detected containing a detection site that has been mutated and an inhibitory polynucleotide complementary to a sequence not to be detected containing the detection site that is unmutated are added to a sample containing a DNA to be detected in which the detection site has been mutated and a DNA not to be detected in which the detection site is unmutated, so that the detection probe is hybridized with the DNA. Then while the hybridization product between the DNA and the detection probe is heated, a signal variation associated with an increase in temperature is measured, then the signal variation is analyzed, and thereby a Tm value is determined, based on which the presence of the mutation is determined.

The invention provides a coronavirus separated from pangolin. The coronavirus is named as pangolin coronavirus xCoV, the homology of the coronavirus xCoV and S protein of SARS-COV-2 reaches 92.5%, receptors of xCoV infected cells and the SARS-COV-2 are consistent and are angiotensin converting enzyme 2 (ACE2). However, the xCoV does not infect people, is very safe to people, can be used for screening active drugs and vaccines for resisting the SARS-COV-2 virus, and can also be used for preparing attenuated vaccines or inactivated vaccines for resisting the SARS-COV-2 virus. Based on the pangolin coronavirus xCoV, a plurality of active compounds with anti-coronavirus activity are obtained by screening, EC50, CC50 and SI evaluation is performed on cepharanthine (stephanine), silamectin and mefloquine hydrochloride (mefloquine) in the active compounds, and the xCoV inhibition mechanism of cepharanthine is analyzed and researched through transcriptome sequencing.

The invention provides a biological repair tablet for endocranium and a preparation method thereof. The repair tablet uses small intestine submucosa tissues of inbred line animals without cell and DNA (deoxyribonucleic acid) components as the raw material, completely reserves the extracellular matrix component and structure, and has a micropore structure. The preparation method of the repair tablet comprises the following operation steps: determination of animal source, pretreatment and rough cleaning of small intestine tissues, virus inactivation, cell removal, DNA removal treatment, formation, packaging and sterilization. The biological repair tablet for endocranium prepared by the method uses an inbred line animal as an animal source, and thus, the hereditary features are pure, stable and uniform, thereby radically ensuring the stability and uniformity of different batches of products; and the biological repair tablet for endocranium has fewer animal source DNA residues, completely reserves the three-dimensional structure of natural ECM, and has the advantages of low immune source property and high infection resistance.

The invention relates to an improved CTAB method for extracting polygonum capitatum DNA. The method comprises the following steps: fetching polygonum capitatum, grinding polygonum capitatum into powder in liquid nitrogen, transferring the powder to a centrifuge tube, adding a CTAB extracting buffering solution, fully shaking the centrifuge tube, carrying out water bath and centrifugation; obtaining the supernate, adding chloroform and isoamyl alcohol with an equal volume, transferring the supernate to another centrifuge tube, collecting the supernate again, extracting the supernate by chloroform and isoamyl alcohol; adding NaAc and isopropanol into supernate, slightly turning over the centrifuge upside down to evenly mix the solution, carrying out centrifugation, discarding the supernate, washing the precipitate by ethanol (70%), carrying out centrifugation, absorbing ethanol by a small gun head, drying the precipitate in the air; removing the ethanol in precipitate, dissolving and digesting the precipitate by TE containing RNA enzymes until the RNA is completely degraded; extracting the solution by phenol, chloroform, and isoamyl alcohol with an equal volume respectively, adding supernate into NaAc and cold anhydrous ethanol, slightly and evenly mixing, carrying out centrifugation, washing the precipitate by an ethanol solution (70%), drying the precipitate in the air, adding a TD solution to dissolve the precipitate, and finally preserving the solution at a low temperature. The modified CTAB method is used to extract polygonum capitatum DNA and has the advantages of simpleness, low cost, and higher purity of polygonum capitatum DNA.

The invention provides a maize gene ZmGRAS37 for regulating and controlling the nutrient body largeness, the early blossoming and the grain weight increment of arabidopsis thaliana at a seedling stage and application thereof. According to the maize gene ZmGRAS37, the gene ZmGRAS37 is separated and cloned from maize; the cDNA (complementary Deoxyribonucleic Acid) sequence of the ZmGRAS37 is connected to an expression vector pPZP211 promoted by a 35S promoter; the arabidopsis thaliana is infected and converted by utilizing an agrobacterium infection method; indicated by an experimental result, the gene can be used for enabling the model-plant arabidopsis thaliana to generate characters of the nutrient body largeness and to further have the early blossoming, the grain weight increment and the like at the seedling stage. Shown by a phylogenetic-tree analysis result, the direct homologous gene of the gene does not exist in the model-plant arabidopsis thaliana; however, the gene has the higher homology with that of sorghum and maize of gramineous plants; the gene is accounted to be the peculiar GRAS (Generally Recognized as Safe) family gene of the graminaceous plants; in consideration of the conservatism of the gene in the graminaceous plants and the phenotypes presented in transgenic arabidopsis thaliana, the gene is proven to have potential application value.

A humanized antibody against tissue factor (TF), comprising:

A: a humanized H chain comprising (1) a H chain V region comprising the H chain CDRs of a mouse monoclonal antibody against TF and the H chain FRs of a human antibody, and (2) the H chain C region of a human antibody; and

B: a humanized L chain comprising (1) an L chain V region comprising the L chain CDRs of a mouse monoclonal antibody against TF and the L chain FRs of a human antibody, and (2) the L chain C region of a human antibody.

After generating a humanized V region by grafting the CDRs of a mouse monoclonal antibody onto a human antibody, a humanized antibody having a higher activity is searched by replacing the FR of the above region with the corresponding FR of the human antibody having a high homology.

The chemically synthesized HSV1 viral gB glycoprotein extracellular region gene fragment and its expression and application relate to the fields of genetic engineering technology, vaccines and diagnostic reagents. The present invention screens out the strong epitope in the gB glycoprotein of HSV1 virus through computer analysis, from the first amino acid to the 696th amino acid, a total of 696 amino acids, selects codons favored by both eukaryotic and prokaryotic organisms, and chemically synthesizes The brand-new gene sequence of the antigenic epitope uses genetic engineering technology to express the gene fragment and prepare a strong antigenic epitope fragment of the gB glycoprotein of the HSV1 virus. The expressed strong antigenic epitope fragment of gB glycoprotein of HSV1 virus can be used for the detection of vaccine, HSV1 virus antibody or antigen, and for immunization preparation of anti-HSV1 virus monoclonal antibody and polyclonal antibody and the like.

The invention discloses a biodegradable collagen-based cornea substitute and a preparation method thereof. The substitute is obtained as follows: collagen and a macromolecular polymer are subjected tocompound crosslinking treatment in a aqueous phase and then cured by a mol; or the collagen and the macromolecular polymer are subjected to electrospinning to form a nanofiber membrane, the nanofibermembrane is further subjected to compound crosslinking and drying to form a 3D cornea scaffold, and the 3D cornea scaffold is used after rehydration swelling clinically. The cornea substitute has controllable degradability, good phototropism and mechanism strength and significant induction functions on increase of corneal epithelial cells and corneal corpuscles.