144 results about "Deoxynupharidine" patented technology

The invention belongs to the technical field of biological medicine, and particularly discloses a composite vaccine adjuvant as well as a preparation method and application thereof. The composite vaccine adjuvant provided by the invention is an oil-in-water emulsion, and simultaneously contains squalene, polysorbate 80, sorbitan trioleate, polyinosinic acid (Poly I: C) and CpG oligodeoxynucleotide (CpG-ODN). The sequence-optimized CpG-ODN and Poly I: C are used in a specific emulsion system in a combined mode, a remarkable synergistic immune enhancement effect can be generated, the induced humoral immunity and cellular immunity response level is remarkably higher than the sum of the effects when all the components are independently used, and the composite adjuvant is good in stability, high in immunostimulatory activity and suitable for clinical application. The immune effect can be remarkably improved by combined application with various vaccine antigens. The invention also provides a preparation method and application of the composite vaccine adjuvant.

The present invention is directed to terminal deoxynucleotidyltransferase (TdT) variants that (i) comprise an amino acid sequence that is at least a specified percent identical to an indicated SEQ ID NOs and have at least one substitution at Q455 or at least Q455 plus at least one further substitution at G186, S248, T331, Q390, K394 or H466 (where positions are with respect to SEQ ID NO 1 and functionally equivalent positions in indicated SEQ ID NOs), (ii) are capable of template-free extension of a polynucleotide, and (iii) exhibit enhanced stability or enhanced efficiency in incorporating 3′-0-blocked nucleoside triphosphates into a polynucleotide. The invention is also directed to the use of these TdT variants for synthesizing polynucleotides of any predetermined sequence.

The present invention relates to methods for producing a protected deoxyribonucleic acid (DNA) product with enhanced resistance to nuclease digestion. The methods comprise digesting a double-stranded DNA molecule with an endonuclease to generate a digested double-stranded DNA molecule comprising a first strand and a second strand, wherein the first strand has a 5' overhang; and extending the second strand 5' to 3' using a polymerase in the presence of one or more nuclease-resistant deoxynucleotide triphosphates to generate the protected DNA product, wherein the protected DNA product comprises an extended second strand comprising at least one nuclease-resistant nucleotide incorporated into the extended second strand by the polymerase using the 5' overhang of the first strand as template. Also provided are protected DNA products and uses thereof.

This invention discloses a method for the bioenzymatic synthesis of deoxynucleoside monophosphate. The invention provides a mutant protein of dNK, comprising mutating amino acid residues at positions 84, 88, 110, and 172 of the amino acid sequence shown in SEQ ID NO:1, while leaving other amino acid residues unchanged, to obtain a protein with deoxynucleoside kinase activity. This invention obtains a deoxynucleoside kinase mutant capable of stably and efficiently catalyzing the synthesis of 3'-O-NH2-deoxynucleoside monophosphate using 3'-O-NH2-deoxynucleoside as a substrate, providing a new approach and pathway for nucleotide production. The bioenzymatic synthesis of (deoxy)nucleoside monophosphate of this invention has advantages such as high conversion rate, mild reaction conditions, simple production process, environmental friendliness, and ease of large-scale production.

The invention discloses a method for synthesizing deoxynucleoside monophosphate by a biological enzyme method. The mutant protein of dNK provided by the invention is a protein with deoxynucleoside kinase activity, which is obtained by mutating amino acid residues at 84th, 88th, 110th and 172nd positions on an amino acid sequence as shown in SEQ ID NO: 1 and keeping amino acid residues at other positions unchanged. According to the present invention, the deoxynucleoside kinase mutant is obtained, 3 '-O-NH2-deoxynucleoside can be adopted as the substrate, the 3'-O-NH2-deoxynucleoside monophosphate can be stably and efficiently catalyzed and synthesized, and the new thought and the new approach are provided for nucleotide production. The (deoxy) nucleoside monophosphate synthesized by the biological enzyme method has the advantages of high conversion rate, mild reaction conditions, simple production process, environmental friendliness, easiness in large-scale production and the like.

This application discloses a linker and its preparation method. The linker includes pharmacophore P, pharmacophore Q, and deoxynucleotides and / or deoxynucleotide derivatives connecting pharmacophore P and pharmacophore Q. Pharmacophore P is obtained by removing the protecting group from group B, and pharmacophore Q is obtained by removing the protecting group from group X. Group B is obtained by chemically modifying the benzylquinolone carboxylic acid pharmacophore, and group X is obtained by chemically modifying the zenomeprazole pharmacophore. The linker is prepared by linking group B and group X with deoxynucleotides and / or deoxynucleotide derivatives to form the linker, including solid-phase synthesis technology. The method of this application can automatically synthesize molecular probes or bivalent drugs containing two pharmacophores, thereby replacing the traditional stepwise linking method, enabling efficient construction of screening libraries, and allowing control of the spatial distance and spatial orientation of the pharmacophores by adjusting the number and base arrangement of deoxynucleotides.

The invention belongs to the technical field of antibody preparation, and particularly relates to an anti-terminal deoxynucleotidyl transferase (TdT) antibody and application thereof. The amino acid sequences of CDR1-3 on a light chain variable region of the antibody are respectively shown as SEQ ID NO.3-5, and the amino acid sequences of CDR1-3 on a heavy chain variable region of the antibody are respectively shown as SEQ ID NO.8-10. The antibody disclosed by the invention can specifically recognize TdT protein naturally expressed in cells and tissues, has the advantages of high specificity, high affinity, high sensitivity and the like when being used for immunoblotting, immunohistochemistry and other immunodetection systems, is beneficial to reducing false positive and false negative rates in pathological diagnosis, improves the accuracy of immunodiagnosis, and has a good application prospect. The method has wide application potential in the field of immunological detection of living biological samples. In addition, the antibody disclosed by the invention has cross-species reaction characteristics on homologous TdT proteins of human, mice and rats, so that the application scene of the antibody is greatly expanded.

Disclosed herein is a composition comprising a lipid nanoparticle and a double-stranded oligodeoxynucleotide (dsODN) encapsulated in the lipid nanoparticle. According to the embodiments of the present disclosure, the dsODN comprises two strands complementary to each other, in which the first strand comprises the nucleotide sequence of SEQ ID NO: 1. Also disclosed herein are methods of treating cancers by using the present composition.

In one aspect, a method of treating an NK cell to induce a gene edit. The method may include contacting an NK cell with a transfection composition including poly-L-glutamic acid (y- PGA), a single-stranded oligo deoxynucleotide (ssODN) enhancer, or a combination thereof; contacting the NK cell with an editing composition including: a DNA-PK inhibitor, and an HD AC inhibitor; electroporating the NK cell; and contacting the NK cell with a recovery composition including DNase I. In another aspect, a method of preparing an engineered NK cell comprising a fluorescent reporter at an endogenous locus. In another aspect, a method of treating an NK cell to introduce an exogenous nucleic acid.

Compounds and pharmaceutical compositions useful to treat cancer (e.g., lymphoma), neurodegenerative diseases, and autoimmune disorders include (1) a first nucleic acid sequence capable of binding to a transcription factor, for example, a signal transducer and activator of transcription (STAT) factor, such as STAT3, (2) a second nucleic acid sequence capable of binding a Toll-like receptor protein, for example, a CpG oligodeoxynucleotide, and (3) a ubiquitin ligase binding compound capable of binding a ubiquitin ligase protein, for example, a compound that is capable of binding a cereblon protein, such as lenalidomide, pomalidomide, or thalidomide.

The invention relates to a high-flux enzymatic DNA synthesis method based on framework nucleic acid. Specifically, the invention provides a TdT-framework nucleic acid compound which can accurately regulate and control the position and distance of terminal deoxynucleotidyl transferase (TdT) so as to achieve the purpose of high-flux enzymatic DNA synthesis. The invention also provides a method for preparing the TdT-framework nucleic acid compound, and a method for synthesizing DNA (Deoxyribonucleic Acid) by using the TdT-framework nucleic acid compound. When the TdT-framework nucleic acid compound is used for synthesizing DNA, high-flux enzymatic DNA synthesis can be realized, and the TdT-framework nucleic acid compound has important significance for developing a high-flux DNA synthesis technology with mild conditions.

Multiple polynucleotides having different, arbitrary sequences are synthesized on the surface of an array by spatial control of polymerase activity. The polymerase is a template-independent polymerase such as terminal deoxynucleotidyl transferase (TdT). Spatial control of polymerase activity is implemented by localized changes in redox-pH conditions. A single species of nucleotide is added and incorporated on growing polynucleotide strands at locations on the array where the polymerase is active. A washing step removes the polymerase and free nucleotides. This process may be repeated multiple times changing both the location of polymerase activity and the species of nucleotide thereby synthesizing different polynucleotides in parallel on the surface of the array. Polymerase activity may be regulated by removing a blocking group attached to a His-tag sequence on the polymerase, a change in pH, or release of encapsulated inhibitors.

The application discloses CpG-containing oligodeoxynucleotide (ODN) with the function of regulating the body's immune capacity and application thereof. The oligodeoxynucleotide in the application is different from the design concept of traditional A-type CpG ODN in sequence and structure, embodies the complex drug-making rule and overcomes the problem of traditional A-type CPG in drug-making. The oligodeoxynucleotide has excellent activity of regulating the body's immune function in vitro and in vivo, can stimulate the production of I, II and III type interferons, regulates the Th1 type immune response, and inhibits the Th2 type and Th17 type inflammatory response. Therefore, the oligodeoxynucleotide can prevent and treat allergic diseases caused by Th2 type immune response, including allergic rhinitis, atopic dermatitis, asthma, chronic obstructive pulmonary disease, eosinophil and Th17 related diseases, can prevent and treat the infection and spread of respiratory or non-respiratory pathogenic microorganisms including viruses, bacteria, fungi and parasites, can improve the immune level of the elderly and immunodeficient population, can convert "cold tumor" into "hot tumor", improve the effect of cancer immunotherapy, can prevent and treat central nervous system diseases related to immune dysfunction such as Alzheimer's disease, and can be applied to other diseases related to immune dysfunction.

The application relates to the technical field of biological medicine, and particularly discloses CpG oligodeoxynucleotides and application thereof. The CpG oligodeoxynucleotide has a sequence as shown in any one of SEQ ID NO. 3-6, 9, and preferably has a sequence as shown in SEQ ID NO. 3-6. The CpG oligodeoxynucleotide can effectively activate the immune system of a human body or an animal, has the functions of preventing and treating pathogenic microorganism infection in a broad spectrum, has the function of improving the immunogenicity of a common vaccine, and can be applied as a human or animal vaccine adjuvant.

The application relates to the field of organic membrane separation, and specifically discloses a method for replacing deoxyribonucleotide triphosphate amine salt with sodium salt by adopting nanofiltration roll type membrane. The method specifically comprises the following steps: a concentration step, concentrating deoxyribonucleotide triphosphate amine salt fraction after high-pressure liquid phase purification; a replacement step, adding 1.5mol / L-6mol / L sodium (potassium) carbonate or sodium (potassium) hydrogen carbonate 0.5V-2V salt solution to the deoxyribonucleotide triphosphate amine salt solution for low-temperature replacement; and a desalination step, introducing carbon dioxide into the system in the above step, and washing with water until sodium ions cannot be detected in the filtered liquid, so as to obtain a deoxyribonucleotide triphosphate sodium salt solution. The preparation method can replace the organic amine salt with sodium salt, and 100% replacement of the organic amine salt can be realized.

A pharmaceutical composition comprising: i) a herpes gE protein, an active fragment of the protein, a variant of the protein, or a mixture of at least two of them; ii) an immunostimulatory composition comprising a saponin and a CpG oligodeoxynucleotide, or consisting of an adjuvant comprising a saponin and a CpG oligodeoxynucleotide. Use of the pharmaceutical composition in the preparation of a medicament for preventing and / or treating a varicella-zoster virus infection and / or a varicella-zoster virus mediated disease. The pharmaceutical composition achieves an unexpected technical effect and can mediate a stronger immune response.

Methods, devices, and compositions are provided for inkjet assisted synthesis of a plurality of polynucleotides at a reaction site on a substrate using a template-free polymerase (e.g., a terminal deoxynucleotidyl transferase (TdT)). Compositions include printable stable formulations of synthetic reagents for inkjet delivery, including, but not limited to, template-free polymerases, divalent cations, and nucleotides.

The application discloses a TdT signal amplification PAM-independent Cas12a lateral chromatography nucleic acid detection method and application. The method comprises the following steps: performing a first reaction on a first reaction system comprising a to-be-detected nucleic acid, a recombinase polymerase amplification reaction component, a recombinase polymerase amplification primer pair and an exonuclease, so as to obtain a single-stranded target nucleic acid; performing a second reaction on a second reaction system comprising the single-stranded target nucleic acid, a CRISPR / Cas12a reaction component, a reporter probe, a terminal deoxynucleotidyl transferase and a deoxynucleotide triphosphate substrate, so as to obtain an extension product; and contacting and hybridizing the extension product with a gold nanoparticle-labeled detection probe in a lateral chromatography process, and when a visual detection signal is obtained, it is judged that the to-be-detected nucleic acid is a target nucleic acid. The method of the application can significantly improve the detection sensitivity and signal contrast while getting rid of the PAM restriction, the detection process does not need complex instruments, the result can be judged by naked eyes, and the method is suitable for on-site rapid visual detection of pathogenic microorganism nucleic acid.

The present invention provides an engineered terminal deoxynucleotidyl transferase (TdT) useful in template-independent polynucleotide synthesis, the terminal deoxynucleotidyl transferase having increased thermostability and / or increased TdT activity and / or increased protein solubility, as well as compositions comprising the said terminal deoxynucleotidyl transferase and methods comprising the use of said terminal deoxynucleotidyl transferase.

This application relates to mutant polymerases. This application discloses mutant phage-type RNA polymerases, such as a mutant T7, SP6, and T3 RNA polymerase, may use 2′-modified nucleoside triphosphates or deoxynucleotide triphosphates as substrates. Methods for producing nucleic acid molecules using these mutant polymerases are also disclosed.

A single-stranded oligonucleotide having a length of 16-30 nucleotides. The single-stranded oligonucleotide and complement component C3 (CC3) mRNA have sufficient complementarity to mediate RNAi effect; each nucleotide in the single-stranded oligonucleotide is a modified or unmodified nucleotide, at least one nucleotide in the single-stranded oligonucleotide is a nucleotide X, and the at least one nucleotide is a fluoro-modified nucleotide; moreover, according to a direction from the 5' end to the 3' end, at least one nucleotide X is located after an eighth nucleotide and is spaced apart from the eighth nucleotide in the single-stranded oligonucleotide by 4-7 nucleotides; and each nucleotide X is a deoxynucleotide or an unmodified nucleotide. The present invention further relates to a double-stranded oligonucleotide comprising the single-stranded oligonucleotide as an antisense strand, an oligonucleotide conjugate, and a pharmaceutical composition.

A comprehensive teaching aid system for genetic science at least including numerous (deoxy-) ribonucleotide models that can be assembled to form DNA / RNA single strands, or to form the beautiful DNA double helix structure when numerous adjacently and oppositely connected deoxynucleotide models are attached by magnets, tRNAs, and three different plates for DNA replication, mRNA transcription and protein synthesis respectively. The (deoxy-) nucleotide model includes a phosphate model, a (deoxy-) ribose model and a base model connected in sequence. Between two adjacently disposed (deoxy-) ribonucleotide models, a (deoxy-) ribose model is connected to a phosphate model in the head-to-tail fashion to form a detachable and flexible chain structure. The base model is laterally connected to the (deoxy-) ribose model, and two base models in two oppositely disposed deoxynucleotide models are flexibly and complementarily attached.

The application belongs to the technical field of pharmaceutical preparations, and discloses a nano-liposome with synergistic effect of double adjuvants and a preparation method and application thereof.The preparation method of the nano-liposome with synergistic effect of double adjuvants comprises the following steps: (1) mixing manganese chloride solution, cationic liposome, distearoyl phosphatidylcholine, distearoyl phosphatidyl ethanolamine-polyethylene glycol, cholesterol and a solvent to react, and evaporating to obtain a lipid film; (2) mixing the lipid film, human unmethylated oligodeoxynucleotide and water to heat, and filtering to obtain the nano-liposome with synergistic effect of double adjuvants.The obtained nano-liposome with synergistic effect of double adjuvants has excellent sustained release property.The obtained nano-liposome with synergistic effect of double adjuvants has stable physicochemical properties after being prepared into a nano-liposome vaccine adjuvant, and is convenient to store and transport.

Provided herein is an antisense oligonucleotide for targeting long non-coding RNA maternally expressed gene 3 (IncRNA Meg3), wherein the oligonucleotide comprises or consists of one of the following sequences in the 5' to 3' direction: (I) C G C dC dG dC dC dA dT dA dT dC dT C C C (SEQ I D NO: 1 ); (ii) C G G dA dC dA dA dA dA dC dT dG dG T T G (SEQ ID NO: 2); (ill) A A G dA dG dT dC dT dC dC dT dC dC dT T A A (SEQ ID NO: 3); (iv) G A T dT dA dG dC dC dC dT dG dT dG dT T C A (SEQ ID NO: 4); (v) C G G dC dA dC dA dA dG dA dG dC dC dA A A G (SEQ ID NO: 5); (vi) T O T dC dC dT dC dC dT dT dA dA dG dC C C C (SEQ ID NO: 6); or (vii) G T C dT dC dC dT dC dC dT dT dA dA dG C C C (SEQ ID NO: 7); wherein dA, dT, dG and dC are deoxyribonucleotides, or wherein dC is 5-methyl-2'-deoxycytidine; and wherein A, T, G and C are modified or unmodified nucleotide building blocks.

The invention provides a method for detecting mycotoxin by using a carbon nanotube field effect transistor biosensor based on nucleic acid conformation change, which comprises the following steps: anchoring a modified nucleic acid aptamer on the surface of an ion shielding gold film layer through a gold-sulfur bond by using a CNT-FET biosensor with an ion shielding gold film layer above a channel to construct a sensing interface; the modified nucleic acid aptamer consists of a target mycotoxin original nucleic acid aptamer sequence and an oligomeric thymine deoxynucleotide fragment directionally inserted into the 5'end of the target mycotoxin original nucleic acid aptamer sequence; during detection, mycotoxin in a to-be-detected sample is specifically combined with the modified nucleic acid aptamer, secondary conformation change of the mycotoxin close to the gold film layer is triggered, the CNT-FET biosensor converts an electric field effect caused by conformation displacement into a channel carrier concentration change signal, and quantitative detection is achieved according to the linear relation between the signal and the mycotoxin concentration. According to the method, the nucleic acid aptamer is directionally modified, so that the electrical response after the low-charge-quantity / electrically neutral mycotoxin is combined with the sensor is enhanced, and the detection sensitivity is greatly improved.

The present disclosure relates to methods for analyzing nucleic acid molecules present in a sample, wherein the DNA molecules present in the sample or derived from nucleic acids in the sample are contacted with a terminal deoxynucleotidyl transferase to add nucleotides to the 3' end of the DNA molecules in a step-wise fashion and then identifying the contacted DNA molecules by their sequence. The method can be advantageously used in methods of diagnosing diseases caused by pathogenic organisms, as well as detecting the presence of pathogenic organisms in the environment.

The invention discloses a vidarabine monophosphate biosynthesis method based on an ATP (adenosine triphosphate) regeneration system, and belongs to the technical field of biomedical engineering. According to the method, riboside arabinoside and ATP serve as reaction substrates, the target product riboside arabinoside monophosphate is synthesized through double-enzyme catalysis, double enzymes are deoxynucleoside kinase and polyphosphate kinase, the deoxynucleoside kinase is mainly used for catalyzing riboside arabinoside to synthesize riboside arabinoside monophosphate, and the polyphosphate kinase is used for catalyzing riboside arabinoside to synthesize riboside arabinoside monophosphate. Polyphosphokinase is mainly used for catalyzing ADP (adenosine diphosphate) and polyphosphoric acid or polyphosphate to regenerate to obtain ATP (adenosine triphosphate). The method can be used for a pure enzyme reaction system, a crude enzyme liquid reaction system or a resting cell catalysis system. The method disclosed by the invention is environment-friendly, realizes low-cost efficient conversion of a substrate through coupled ATP regeneration, plays a significant role in promoting biosynthesis of arabinoside monophosphate series nucleoside drugs, and has application potential and economic value.

The invention provides a triple signal amplification electrochemical method for detecting a porcine reproductive and respiratory syndrome virus NADC30-like strain. Firstly, the surface of a gold electrode is modified with a hairpin signal probe H1, when a target sequence appears, the hairpin H1 is opened, a sequence capable of triggering a hybridization chain reaction (HCR) is exposed, long dsDNA is formed, and first signal amplification is achieved. Then, under the catalysis of terminal deoxynucleotidyl transferase (TdT), a large amount of polythymine (PolyT) extends from 3'terminal hydroxyl groups of H2 and H3 fragments, and second signal amplification is realized; polyT serves as a template, in-situ deposition of a large amount of CuNPs is caused in a short time, the signal response of the electrochemical sensor is remarkably enhanced, and third signal amplification is achieved. And finally, monitoring metal ions released by dissolution of the nanoparticles by using a differential pulse voltammetry, and drawing a standard curve through current change to realize accurate quantification of the target virus. The method is high in sensitivity and good in specificity, and can be used for general detection of viruses by replacing a target virus specific sequence and a corresponding probe.

The application discloses a composite adjuvant containing CpG oligodeoxynucleotide and aluminum adjuvant, and a new coronavirus COVID-19 inactivated vaccine using the composite adjuvant. The CpG oligodeoxynucleotide is used in combination with the aluminum adjuvant, and a strong immune enhancement effect can be generated. When the composite adjuvant is applied to the new coronavirus COVID-19 inactivated vaccine, the residence time of the antigen in the body can be greatly prolonged, and the hydrolysis of the antigen by various hydrolytic enzymes is reduced. The preparation method of the new coronavirus COVID-19 inactivated vaccine provided by the application is simple, the quality is easy to control, the vaccine is easy to mass-produce, has good safety, low toxic and side effects, and is particularly suitable for being used for the prevention and treatment of the new coronavirus COVID-19 in various populations, including the low-immunity population such as the middle-aged and the old.