138 results about "Luciferases" patented technology

The invention relates to the technical field of bacterial detection, in particular to a bacterial drug sensitivity detection assembly based on an ATP luciferase method and a using method.The bacterial drug sensitivity detection assembly comprises a supporting assembly, a sampling assembly, an ATP fluorescence detector and a 96-hole drug sensitivity plate, and a sealing block is arranged at the top of the inner wall of the ATP fluorescence detector; the supporting assembly comprises a base, a controller is arranged on one side of the base, and an electric push rod is arranged in the middle of the upper surface of the base. The device effectively realizes detection of bacterial drug sensitivity of a sample, is high in detection precision and good in detection effect, can also mix and stir the sample in the 96-hole drug sensitive plate, and avoids precipitation of bacteria in the 96-hole drug sensitive plate due to long-time standing; the sampling quantity of samples can be accurately detected, and the accuracy and the stability of subsequent detection are ensured; when the sample in the sampling tube is mixed with the reaction liquid, the flowing efficiency of the sample can be ensured, and the problems of high liquid feeding difficulty, poor mixing effect and the like caused by negative pressure are avoided.

The invention provides a chicken PGC reporter gene cell strain for detecting the biological activity of FGF2 as well as a construction method and application of the chicken PGC reporter gene cell strain, and belongs to the technical field of biological activity detection. The cell strain is a recombinant cell which is stably integrated with a reporter gene expression cassette responding to an FGF signal channel in a chicken PGC genome; the expression cassette comprises a serum response element (SRE), a minimum promoter and a reporter gene (such as a luciferase gene). The construction method comprises the following steps: introducing a donor vector containing the elements into chicken PGC by utilizing a PiggyBac transposon system, and screening to obtain a stably expressed cell strain. The cell strain provided by the invention retains endogenous signal channel characteristics of chicken PGC, and can specifically and highly sensitively respond to FGF2 stimulation. The method can be used for biological activity quantitative determination and thermal stability evaluation of the FGF2 protein and the mutant thereof, quality control of culture medium components and screening of an FGF signal channel regulator.

A plasmid capable of automatically inserting a membrane-localized luciferase reporter gene into a chromosome of a bacterium / recipient bacterium. After the plasmid inserts a luciferase reporter gene into a chromosome of a bacterium / recipient bacterium, the plasmid can be lost. Since the plasmid can be lost, the bacterium / recipient bacterium into whose chromosome the luciferase reporter gene is inserted does not require the addition of antibiotics during subsequent use, providing convenience and reducing interference.

The invention provides a single-chain and double-protein-expressed circular RNA construct and a preparation method and application thereof, and the single-chain and double-protein-expressed circular RNA construct comprises 5'and 3 'introns used for realizing RNA self-splicing and cyclization as a first kind of introns; the 5'and 3 'homologous arms are used as pairing sites in RNA molecules; the exon 1 and the exon 2, which are connected after being spliced, can be spliced and connected together under the action of the first type of introns to form a complete and continuous open reading frame; an interval sequence for providing spatial and adjustment sequence structures; the IRES is used for starting translation; a glycine-serine linker as a flexible linker; a green fluorescent protein for gene expression monitoring; a luciferase for producing light by a catalytic chemical reaction. All the advantages of the double-IRES system for generating natural complete protein are reserved, and the core defect of low efficiency can be thoroughly overcome.

The present invention relates to novel cell lines for the expression of the human LHCG receptor and for the expression of the human LHCG receptor and a luciferase reporter gene. Methods for producing such cell lines are also described. The cell lines of the present invention, which express the LHCG receptor and a luciferase reporter gene, can be used in methods for determining the presence and biological activity of r-hLH and r-hCG samples.

The present invention relates to a method for screening compounds for bactericidal activity using thermostable luciferase and based on a real-time bioluminescence measurement. The present invention further relates to a method for determining the sensitivity of a bacterial sample originating from a subject suffering from a bacterial infection to a group of known antibiotics and to a method for assessing the minimum inhibitory concentration (MIC) of a bactericidal compound.

The present disclosure provides polypeptides that can be used in a complementation assay as a biosensor. In certain aspects, the polypeptide is a larger component of a split luciferase, wherein the luciferase is derived from LuxSit. The polypeptide may comprise an amino acid sequence having at least 85% identity to the amino acid sequence: of SEQ ID NO:1 and comprises D at position 30, N at position 46, R at position 58, R at position 61, F at position 64, T at position 68, E at position 101, and A at position 103; G at position 9, D at position 30, D at position 54, F at position 64, K at position 68, and E at position 101; D at position 30, D at position 46, K at position 68, and E at position 101; S at position 9, D at position 30, L at position 43, D at position 46, G at position 54, F at position 64, K at position 68, E at position 101, and T at position 103; D at position 30, D at position 46, F at position 64, K at position 68, E at position 101, and T at position 103; N at position 30, D at position 46, E at position 101, and A at position 103; or D at position 30, N at position 46, F at position 64, T at position 68, E at position 101, and A at position 103, wherein the numbering of the amino acid positions is based on the numbering of the amino acids in SEQ ID NO:1.

The present disclosure is directed to compositions comprising mitochondrial optogenetics-based gene therapies and their methods of use. In some embodiments, a composition described herein comprises an expression vector comprising a first nucleic acid sequence encoding a channelrhodopsin fusion protein and a second nucleic acid sequence encoding a luciferase protein. In some cases, the first nucleic acid sequence and the second nucleic acid sequence are operably linked to an expression control sequence. In some instances, the channelrhodopsin fusion protein comprises a channelrhodopsin protein linked to an inner mitochondrial membrane-mitochondrial localization signal (IMM-MLS). In some implementations, when the expression vector is expressed, the luciferase protein is localized to the cytosol. In some cases, the IMM-MLS comprises a leading sequence from a mitochondrial inner membrane protein selected from ABCB10, ABCB140, Cytochrome C, and renal outer medullary potassium channel (ROMK).

This invention pertains to modern agriculture, specifically the application of transcription factors CdZF-HD1 / 9 in regulating squalene synthesis. Using the CdSQS promoter as bait, this invention screens a yeast single-hybrid library to identify ZF-HD transcription factors that interact with the CdSQS promoter. ZF‑HD1 and ZF‑HD9 .clone ZF‑HD1 and ZF‑HD9 The nucleotide sequence was further confirmed by dual-luciferase assay and EMSA. ZF‑HD1 and ZF‑HD9 Interacts with CdSQS. ZF‑HD1 and ZF‑HD9 Genetic transformation of Camellia oleifera from Vietnam resulted in increased expression levels of squalene synthase and higher squalene content in the induced hairy roots. This invention provides a scientific basis for finding optimal genetic modification and exogenous regulation methods to increase squalene content in Camellia oleifera, and offers important theoretical and practical support for promoting the development of the Camellia oleifera industry in Hainan.

The invention discloses a recombinant foot-and-mouth disease virus carrying a HiBiT luciferase reporter gene as well as a construction method and application of the recombinant foot-and-mouth disease virus. According to the invention, based on a reverse genetics technology, on the basis of full-length cDNA infectious clone of an FMDV O / BY / CHA / 2010 strain, a recombinant foot-and-mouth disease virus carrying a HiBiT luciferase reporter gene is constructed, and the recombinant foot-and-mouth disease virus is successfully rescued in a BSR / T7 cell. Experimental results show that the recombinant foot-and-mouth disease virus has no significant difference from parent viruses in the aspects of replication ability and pathogenicity, and good biological characteristics are maintained. Meanwhile, a cell line capable of stably expressing the LgBiT protein is successfully constructed, real-time detection of light-emitting signals in the virus infection process is achieved through specific binding of the LgBiT and the HiBiT protein, and an effective technical means is provided for virus research and rapid detection.

This invention relates to an antibacterial membrane-penetrating peptide and its applications. The amino acid sequence of the antibacterial membrane-penetrating peptide includes the sequence shown in SEQ ID NO.1 or a sequence with more than 70% homology to the sequence shown in SEQ ID NO.1. This invention screened and obtained a short peptide that specifically binds to the bacterial Hsp70s protein. It was found that this peptide binds to the Hsp70s protein through a traditional substrate-binding pocket, exhibiting an unusually high affinity for the protein in the DnaK-ATP state. The short peptide can inhibit Hsp70s protein activity to a certain extent, preventing it from aiding in the renaturation of denatured luciferase. Furthermore, while maintaining the integrity of the bacterial cell membrane, this short peptide has a superior ability to cross the bacterial cell membrane. The invention further discusses its application potential in the preparation of antibacterial drugs and antibiotic adjuvants, providing new ideas for solving current problems such as bacterial resistance and the difficulty in discovering new antibiotics.

The invention discloses a luciferase mutant capable of efficiently utilizing non-natural nucleoside triphosphate. In the mutant, histidine (H) at the 244 site is mutated into tryptophan (W) on the basis of a starting enzyme Luc146-1H2 (K357E) at which amino acid No.357 is mutated from lysine (K) to glutamic acid (E). Compared with an evolutionary starting enzyme, the mutant Luc-WE not only can more efficiently utilize ATP (adenosine triphosphate) as an energy donor, but also can more efficiently utilize non-natural nucleoside triphosphate as an energy donor to catalyze the oxidation of D-fluorescein, so that a luminescence value higher than that of the evolutionary starting enzyme is generated; for different nucleoside triphosphate energy donors, the luminescence value is 2-40 times that of an evolution starting enzyme. Under the reaction condition of lower nucleoside triphosphate concentration, the luciferase mutant Luc-WE shows higher sensitivity, and has wider application range and practical value.

The present invention belongs to the technical field of genetic engineering, and specifically relates to a copepod luciferase mutant and the use thereof. The mutant is obtained by means of performing one or more mutations on a wild-type copepod luciferase with the signal peptide removed, and can be expressed in a prokaryotic expression system, resulting in an increased luminescence half-life or a signal rise within a certain time range, and the luminescence pattern of the luciferase is converted to glow-type luminescence or luminescence with a sustained increase in luminescence intensity. Some copepod luciferase mutants exhibit enhanced luminescence intensity. The mutant is widely used in biological research for real-time tracking and continuous monitoring, such as for labeling and tracking activities or processes in living organisms. On the basis of the advantage of increased luminescence half-life of the copepod luciferase mutant, the mutant can be used as and / or in the preparation of a glow-type luciferase. The copepod luciferase mutant is more suitable as a reporter gene for in-vivo detection in scientific research, and is conjugated with avidin, a digoxin antibody, etc., for sequencing, thereby shortening the sequencing time and improving the sequencing quality.

The invention discloses an application of a CTL-S1 gene and miR-let-7 in regulation and control of reproductive development of bactrocera cucurbitae. The CTL-S1 gene has high specificity expression in bactrocera cucurbitae ovary, and the hatching rate of offspring can be reduced by inhibiting the expression of the CTL-S1 gene through RNA interference. Through the analysis of a dual luciferase report system of RNA co-immunoprecipitation, the result shows that the miR-let-7 and the CTL-S1 have a targeting relationship. Overexpression of miR-let-7 can lead to significant reduction of the expression quantity of the CTL-S1 gene, and a phenotype similar to RNA interference inhibition of the expression of the CTL-S1 gene appears, so that the hatching rate of offspring is reduced. The invention provides a potential gene target for developing a genetic control strategy for destroying the reproduction of bactrocera cucurbitae, provides a new thought for the subsequent biological control of insects by adopting a sterile technology, and has a wide application prospect.

The application discloses a chrysanthemum constitutive high-expression promoter and application thereof in gene editing. The nucleotide sequence of the promoter CmUbi is shown as SEQ ID NO. 4. Through double luciferase reporter experiments and stable genetic transformation experiments, it is proved that the expression activity of the CmUbi promoter in chrysanthemum protoplast and callus is significantly higher than that of commonly used CaMV 35S, AtRPS5a and corn ZmUbi promoters. A CRISPR / Cas9 gene editing vector is constructed by using the promoter, and a key gene of branch of hexaploid chrysanthemum is edited BRC1b , and a plurality of allele simultaneous mutation complete editing strains are successfully obtained, and the number of branches of the mutants is significantly increased. The endogenous high-activity promoter of the chrysanthemum provided by the application effectively solves the problems of low expression activity of an existing heterologous promoter in the chrysanthemum and poor gene editing efficiency of a polyploid, and provides an efficient and adaptive expression regulation element for chrysanthemum molecular breeding and functional genomics research.

The application discloses CTL-S1 A gene and an application of miR-let-7 in regulation of reproductive development of drosophila bacterea CTL-S1 The gene is specifically highly expressed in ovaries of drosophila bacterea, and the expression of the gene is inhibited by RNA interference CTL-S1 The expression of the gene can reduce the hatching rate of offspring. Through analysis of an RNA immunoprecipitation double luciferase reporter system, it is found that miR-let-7 and CTL-S1 there is a target relationship. Overexpression of miR-let-7 can cause CTL-S1 the expression amount of the gene to be significantly reduced, and a phenotype similar to the expression of the gene being inhibited by RNA interference CTL-S1 CTL-S1 CTL-S1 CTL-S1 CTL-S1 CTL-S1 CTL-S1 CTL-S1 CTL-S1 CTL-S1 CTL-S is generated, and the hatching rate of offspring is reduced. The application provides a potential gene target for developing a genetic control strategy for destroying the reproduction of drosophila bacterea, provides a new idea for subsequent biological control of insects by using the sterile technique, and has a wide application prospect.

The application belongs to the field of biology and relates to a nanobody targeting calexcin and a preparation method and application thereof, the nanobody targeting calexcin is successfully screened and prepared, and the nanobody-HRP fusion protein used for affinity identification in an enzyme-linked immunosorbent assay is successfully expressed; in combination with a phage-assisted continuous and discontinuous directed evolution (PACE) system, the nanobody screened is subjected to directed evolution, and stable mutation sites are found through sequence alignment analysis; it is proved through an indirect enzyme-linked immunosorbent assay and a luciferase complementation assay that the affinity of the nanobody after evolution to calexcin is improved by about 2 times.

In one embodiment, the present application provides a liquid circulating luminescent reagent excellent in stability. In one embodiment, the present application provides a liquid composition for measuring ATP, and AMP and / or ADP in a sample after storage, (i) the liquid composition comprising luciferase, luciferin, an enzyme catalyzing a reaction of generating ATP from AMP, a substrate and a co-factor of the enzyme catalyzing the reaction of generating ATP from AMP, or in the case where at least one of these components is not contained in the liquid composition, the component not contained in the liquid composition is added to the liquid composition before or at the time of measurement, and, (ii) the relative luminescence amount of the liquid composition at the time of storage is 5500 RLU or less, the relative luminescence amount being a value after subtracting a control value from a measurement value.

This invention discloses tree fern. AspiMYB5 The application of genes in regulating the biosynthesis of tree fern phenols belongs to the field of genetic engineering technology, which involves regulating the biosynthesis of tree fern phenols within plants. AspiMYB5 The expression level of the gene, thereby regulating the key enzyme gene of tree fern phenol. AspiPKS6 The expression of [a specific enzyme] and the synthesis of tree fern phenol. This invention uses a yeast library screening method to screen genes affecting key enzymes. AspiPKS6 Expressed transcription factors AspiMYB5 The results were verified using yeast single-hybrid technology (Y1H) and electrophoretic mobility variation analysis (EMSA). AspiMYB5 and AspiPKS6 The promoter exhibits direct physical binding in vitro. Dual-luciferase reporter gene assays further demonstrate that... AspiMYB5 and AspiPKS6 When the promoter is co-expressed, the fluorescence signal intensity is significantly reduced, further confirming... AspiMYB5 It can inhibit AspiPKS6 AspiMYB5 Promoter activity. This invention not only improves the theoretical research on the biosynthetic pathway of tree fern phenols, but also provides key target genes and technical support for regulating the content of secondary metabolites in rare plants through genetic improvement.

Compounds that may inhibit Oplophorus-derived luciferases are disclosed as well as compositions and kits comprising the compounds and methods of using the compounds.

The present invention relates to a test kit for performing an assay to detect and measure at least one autoantibody molecule in a subject developing type 1 diabetes (T1D). The invention more specifically relates to a recombinant antigen molecule fused to a bioluminescent reporter protein to detect an autoantibody molecule against a self-antigen. Said self-antigen is preferably expressed by pancreatic islet beta cells and include insulin, proinsulin, Glutamate decarboxylase 65, islet antigen 2, and Zinc transporter 8. More specifically, the present invention relates to an assay comprising a set of at least three recombinant antigen molecules in a single composition, the antigen molecules being selected from the list comprising recombinant human Glutamate decarboxylase 65, recombinant human islet antigen 2, recombinant human Zinc transporter 8, recombinant human insulin, and recombinant human proinsulin. The invention further relates to methods for determining the presence and / or level of at least one autoantibody molecule in a sample from a human subject developing T1D using said recombinant antigen molecules. According to the invention, the presence, absence, or amount of one or more autoantibody molecules against said self-antigens is determined with a luciferase immune precipitation system (LIPS) assay or a modified version of a LIPS assay, i.e., a solid phase capture LIPS (scLIPS) assay.

This invention provides a method for constructing a visualized mouse model characterizing hepatocyte necrosis and apoptosis, belonging to the field of biomedical technology. The invention involves disassembling firefly luciferase (Fluc) into inactive N-terminal (Nluc) and C-terminal (Cluc) fragments, which are then fused and expressed at specific locations on the MLKL molecule, forming two independent plasmid vectors, MLKL-Nluc and MLKL-Cluc. These two plasmid vectors are then injected into mouse hepatocytes via tail vein high-pressure hydrodynamic injection to construct the visualized mouse model characterizing hepatocyte necrosis and apoptosis. This invention allows for non-invasive and dynamic monitoring of the execution process of necrosis and apoptosis—MLKL oligomerization—in animal models, providing a powerful and specific tool that fills the technological gap in real-time visual monitoring of necrosis and apoptosis in pan-apoptotic research.

The application belongs to the field of plant synthetic biology and plant gene expression regulation, and particularly relates to a vector for plant promoter double system quantitative characterization, a standardization method, a product and application. The method combines a protoplast transient expression system and an agrobacterium-mediated leaf transient expression system, uses a compact promoter characterization plasmid (pLIP) containing a firefly luciferase module driven by a to-be-detected promoter and a renilla luciferase internal reference module driven by a 35S promoter, and the two modules are located in a T-DNA interval. The ratio of the two luciferases is calculated, the RPU (relative promoter unit) method is introduced, a standard promoter (35S) is used as a reference, and normalization and direct comparison of promoter activities in different systems are realized. The scheme significantly improves the repeatability and cross-system compatibility of promoter activity data, and is suitable for efficient screening and standardized characterization of regulatory elements in plant synthetic biology.

This invention discloses an attenuated chikungunya virus expressing a nanoluciferase gene, the nucleotide sequence of which is shown in SEQ ID NO:1. It is constructed by attenuating the virulence of the chikungunya virus LR2006-OPY1 strain by deleting 61 amino acids from positions 323 to 383 of the nsP3 protein and inserting a nanoluciferase gene after position 490 of the nsP3 protein. This virus model maintains similar growth characteristics to the wild-type virus but with significantly improved safety. Viral replication can be monitored in real time by detecting luciferase activity, making it suitable for drug screening at both cellular and animal levels. This provides an efficient and safe technical platform for the development of antiviral drugs against chikungunya virus.

The invention discloses a DNA (Deoxyribonucleic Acid) molecule for promoting mRNA (Messenger Ribonucleic Acid) expression and 3'UTR (Untranslated Region) in the mRNA molecule obtained by transcription of the DNA molecule, and the DNA molecule is derived from COX7B of a cytochrome C oxidase family. The invention also discloses an application of the composition containing the 3 'UTR in preparation of RNA (Ribonucleic Acid) therapeutic drugs or mRNA (Messenger Ribonucleic Acid) vaccines. Compared with a control group alpha-globin, the insertion of the 3'UTR fragment provided by the invention has the advantages that the expression quantity of renilla luciferase in Lent-X 293T cells is increased by 2.03 times, and the expression quantity of renilla luciferase in HeLa cells is increased by 3.62 times. The invention shows that the 3 'UTR has an excellent prospect of being applied to RNA (Ribonucleic Acid) therapeutic drugs or mRNA vaccines.

The application discloses a method for identifying Klebsiella pneumoniae gene circadian rhythm expression by using a bioluminescence reporting system. The application constructs a recombinant bioluminescence Klebsiella pneumoniae driven by a metabolic gene malate:quinone oxidoreductase mqo promoter (Pmqo), and directly observes the change rule of bacterial gene expression under entrainment and free-running conditions of temperature cycle by monitoring the luminescence characteristics. The luxCDABE gene operon comprises genes for coding luciferase and substrate synthesis enzyme, wherein luxA and luxB code for heterodimer luciferase, luxC, luxD and luxE synthesize aldehyde as a substrate for the luciferase luminescence reaction. The luciferase catalyzes the oxidation of aldehyde to generate visible blue-green light with a wavelength of 450-490 nm. The Lux bioluminescence reporting system has the advantages of not needing to add an exogenous substrate and not affecting the normal physiological functions of the host, and can be applied to the identification of Klebsiella pneumoniae gene circadian rhythm expression.

The invention provides a compound and application thereof in sequencing. The compound has a structure as shown in a formula (I ') or a stereoisomer, a tautomer or a salt of the stereoisomer, the tautomer or the salt of the stereoisomer, the tautomer or the salt of the stereoisomer, the tautomer or the salt of the stereoisomer, the tautomer or the salt of the stereoisomer, the tautomer or the salt of the stereoisomer. The whole sequencing process is low in cost, and reagents are easy to transport and store.

The invention discloses an oat endogenous U3 promoter AsU3-3p with high transcriptional activity and a preparation method of the oat endogenous U3 promoter. The invention provides a DNA (deoxyribonucleic acid) molecule which is any one of the following: A1) a DNA molecule containing a nucleotide sequence as shown in a sequence 1; a2) a DNA molecule which is obtained by substitution, deletion or addition of one or more nucleotides in A1) and has the same function. According to the invention, 36 U3 promoters are excavated from an oat genome for the first time, and transcriptional activity analysis is carried out on 16 promoters in combination with a dual-luciferase report experiment. The result shows that the oat endogenous U3 promoter AsU3-3p with high transcriptional activity is excavated by analyzing the average luminous intensity and the relative fluorescence intensity of luciferase through Image J, and the oat endogenous U3 promoter AsU3-3p can be used as an ideal promoter for driving sgRNA in an oat CRISPR-Cas system.

The invention provides a biosensor for detecting nitrate content as well as a preparation method and application thereof, and relates to the technical field of biology. The biosensor disclosed by the invention comprises a luciferase amino terminal fragment, a luciferase carboxyl terminal fragment and NasR protein, the luciferase amino terminal fragment and the luciferase carboxyl terminal fragment are two fragments of the same luciferase. The biosensor provided by the invention has high selectivity on nitrate, can specifically, stably, continuously and online observe the nitrate content in the whole plant for a long time, and overcomes the key problems of destructive sampling, limited spatial resolution, insufficient detection sensitivity and the like in the traditional method; and a real-time dynamic detection platform capable of monitoring the whole plant living body is provided for plant nitrogen metabolism research.