21 results about "Luciferase Gene" patented technology

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.

The invention discloses a double-report plasmid system and a construction method and application thereof. The double-report plasmid system comprises a translation report plasmid and an error folding response plasmid; the translation reporter plasmid comprises a target protein gene sequence, an mRNA secondary structure sequence and a luciferase gene sequence; the misfolding response plasmid comprises a heat shock pressure promoter and a fluorescent protein gene. The double-report plasmid system can quantitatively and visually detect translation characteristics and wrong folding conditions of the recombinant protein, the whole operation process is carried out based on a 96-well plate, the double-report plasmid system has the characteristics of low cost, high timeliness, high flux and the like, and the recombinant protein can be rapidly and efficiently soluble (positive: complete soluble; negative: completely insoluble; and in an intermediate state, the solubility is relatively poor), so that the efficiency of recombinant protein expression research is greatly improved.

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 relates to the technical field of gene editing, in particular to a preparation method and application of a monoclonal antibody with a nano luciferase label. A CRISPR / Cas9 gene editing technology is utilized, a nano luciferase gene is knocked into an Ig gene of a hybridoma cell secreting a monoclonal antibody to obtain a stable cell strain with a function of secreting the monoclonal antibody with the nano luciferase label, and then the monoclonal antibody with the nano luciferase label is prepared by utilizing the stable cell strain. According to the method provided by the invention, the sensitivity of an antigen immunoassay method can be improved from the perspective of signal labeling, and the detection limit is reduced.

To provide a luminescence detection method for detecting whether or not a substance such as a drug has reached a target area by detecting luminescence signals from a deep part inside the body producing a non-human animal capable of stably expressing modified luciferase over a long time.SOLUTION: A disclosed luminescence detection method is a method including administering luciferase luminescent substrate and siRNA that suppresses expression of a modified luciferase gene to a transgenic non-human animal carrying modified luciferase genes. The modified luciferase gene expresses modified luciferase that makes a luminescent substrate emit light with higher luminescence intensity than wild-type D-luciferase.SELECTED DRAWING: Figure 5

The invention provides an in-vivo and in-vitro double visual recombinant mouse pox virus ECTVC genome, a recombinant virus, a construction method and application, and belongs to the technical field of gene engineering. The method comprises the following steps: improving a wild type rat poxvirus genome, namely knocking out a non-coding region between C12R and C13R, and inserting a red fluorescent protein gene mKate and a Gaussian luciferase gene Gluc by using a homologous recombination method. The invention also provides a recombinant mouse poxvirus ECTVC containing the recombinant mouse poxvirus ECTVC genome, the obtained recombinant virus has good hereditary stability, and the stability of the recombinant virus can be continuously maintained for at least seven generations. The recombinant virus provided by the invention can be used for in-vitro high-throughput screening of drugs, and can realize visualization of replication and distribution of the virus in a mouse body.

The invention provides a high-sensitivity in-vitro pyrogen detection method, which is characterized in that on the basis of an in-vitro pyrogen detection method (reporter gene method), a transgenic cell which expresses a Toll-like receptor and transfects an NF-kB reaction element and a luciferase gene is used for detecting a pyrogen, the pyrogen is combined with the Toll-like receptor on the surface of the cell to activate an NF-kB signal channel, and the NF-kB reaction element and the luciferase gene are converted into the NF-kB signal channel. Therefore, the expression of luciferase is activated.

The invention relates to a construction method and application of a colorectal cancer liver metastasis cell strain, and the construction method specifically comprises the following steps: transfecting a tdToma / Luciferase gene to CT26. WT to obtain a CT26-TL cell strain; the colorectal cancer cell strain capable of directionally and stably transferring the liver is obtained by taking a CT26-TL cell strain as a starting material and carrying out three rounds of spleen injection screening and five rounds of cecum injection screening, the colorectal cancer cell strain is named as a mouse colorectal cancer cell clone strain CT26-TL-CL5, and the preservation number is CCTCC NO: C2024307. According to the CT26-TL-CL5 cell strain, the technical difficulty of constructing a liver metastasis model starting from in situ in basic research is solved, the problem of observing colorectal cancer liver metastasis in a living body or an isolated organ is solved, and the modeling time is effectively shortened. The CT26-TL-CL5 cell strain can provide a good colorectal cancer liver metastasis animal model for researching molecules and mechanisms for promoting colorectal cancer liver metastasis, and provides cells and related animal models for industrially screening drugs related to colorectal cancer metastasis.

The invention discloses a construction method and application of an asbestos-caused lung carcinoma-in-situ animal model, and belongs to the technical field of biomedicine. The construction method comprises the following steps: firstly, carrying out multiple poisoning treatments on embryo fibroblasts by chrysotile, and then carrying out malignant transformation to obtain malignant transformed cells; a luciferase gene is introduced by utilizing lentivirus transfection; and injecting the successfully transfected malignant transformed cells into the lung of an experimental animal by adopting an intrapulmonary injection method, and feeding to obtain the lung carcinoma-in-situ animal model. The method creatively uses asbestos to induce malignant transformed embryo fibroblasts with cancerous characteristics to establish the lung in-situ cancer animal model, and has the advantages of short modeling period, high tumor formation rate, visualized cell transfer condition, capability of better reflecting the cancerous characteristics of asbestos and the like. The constructed model can be applied to clinical diagnosis and treatment research such as pathogenesis research and drug screening evaluation.

Provided are a fusion antigen of porcine Getah virus (GETV), a kit, a preparation method therefor and an application thereof. The fusion antigen of the GETV is primarily prepared by recombining a Gaussia luciferase (GLuc) gene with a codon- optimized GETV E2 antigen gene to construct an expression vector, and transfecting the expression vector containing GLuc-E2 into mammalian cell lines, resulting in the secretion of GLuc-E2 proteins into a cell supernatant for expression. Without the need for protein purification step, the cell supernatant may be directly collected for disease detection. The present disclosure demonstrates strong specificity and shows no cross- reactivity with African swine fever virus (ASFV), porcine reproductive and respiratory syndrome virus (PRRSV), porcine circovirus type 2 (PCV2), pseudorabies virus (PRV), or Japanese encephalitis virus (JEV).

The present invention relates to novel luciferase genes that exhibit improved characteristics when expressed in cells as a reporter gene and method of using the same for detecting and optionally quantitating the activity of pharmacologically active molecules in a test sample. The invention further relates to a method for detecting and optionally quantitating neutralizing antibodies against a pharmacologically active molecule present in a test sample using the reporter cell line of the present invention. The invention further relates to a method for high-throughput screening in drug discovery for detecting and optimally monitoring biologic processes with optimal sensitivity, signal strength, and biological fidelity using said luciferase genes as reporter genes.

To provide a cancer cell strain that readily forms subcutaneous tumors and facilitates in vitro and in vivo therapeutic testing.SOLUTION: The present invention discloses the following: a cancer cell strain that can form subcutaneous tumors and exhibits RET fusion gene positivity; a cancer cell strain derived from pleural effusion of a human patient with RET fusion gene-positive lung cancer; a cancer cell strain deposited under an accession number NITE P-03936; and a cancer cell strain incorporating an additional luciferase gene.SELECTED DRAWING: Figure 3

The invention discloses a mouse ovarian cancer carboplatin drug-resistant cell strain as well as a construction method and application thereof, and belongs to the field of biomedicine. According to the invention, a firefly luciferase gene is transfected and introduced into a mouse epithelial ovarian cancer ID8 cell strain through a lentiviral vector, and a cell strain (ID8-Luc) capable of stably expressing the luciferase gene is obtained through puromycin screening. In-vitro induction is carried out by gradually increasing the dosage of carboplatin, the carboplatin-resistant ID8-Luc cell strain is successfully constructed, the drug-resistant strength is high, and the drug-resistant stabilization time is long. The invention provides an efficient tool cell strain for exploring a tumor drug resistance mechanism and screening anti-tumor drugs and novel targeted drug-resistant drugs, and has a good application prospect.

The present invention concerns bioluminescent plants and methods for producing them. Among said methods, the invention relates, in particular, to a method of introducing luciferase genes and luciferin biosynthesis genes into the chloroplasts of a plant. In addition, the invention concerns a method of producing light comprising adding chemical compounds to a culture medium of the plant according to the invention.

The invention belongs to the technical field of biology, and particularly relates to a system for improving circular RNA expression in mammalian cells. The construction method of the system plasmid comprises the step of constructing the optimized Tornado system plasmid by sequentially connecting a luciferase gene, a Tornado system cyclization element, a circRNA translation expression cassette and a target gene. According to the present invention, by prolonging the distance between the CMV promoter and the IRES, and increasing the 5 'LTR (including the upstream RSV promoter) and the WPRE, the expression cassette capable of significantly improving the protein expression level of the IRES-containing exogenous circRNA is constructed, and the RNA cyclization system can be used for expressing the exogenous functional target protein so as to be used for the research and development of the therapeutic drug or the prophylactic vaccine.

The invention discloses a cell model for detecting monocyte and macrophage IL-1beta response level by using secretory luciferase, and belongs to the technical field of cell models. The technical problem to be solved is that in the prior art, a method capable of tracking response levels of mononuclear and macrophage factors IL-1beta in a real-time and non-invasive manner under different stimuli is lacked. According to the technical scheme, the cell model for detecting the IL-1beta response level of the mononuclear and macrophages is provided, and a preparation method of the cell model comprises the steps that an IL-1beta promoter is placed on the upstream of a secretory luciferase gene, lentivirus plasmids are introduced, and recombinant lentivirus plasmids are obtained; co-transfecting a host cell with the recombinant lentivirus plasmid and a packaging plasmid to prepare a recombinant lentivirus; and introducing the recombinant lentivirus into the THP-1 cell to obtain the THP-1 cell for expressing luciferase.

The invention discloses a construction method of a herpes simplex virus, and belongs to the technical field of biology. The invention discloses a construction method of a herpes simplex virus. The construction method comprises the following steps: purifying HSV (herpes simplex virus) plaque, extracting a virus genome, and editing a CRISPR-CAS9-mediated TK gene. The gene editing efficiency is remarkably improved through the synergistic effect of the double sgRNAs. The two sgRNAs designed for the HSV-1 thymidine kinase gene respectively target upstream and downstream regions of the TK gene, so that the probability of homologous directional repair is greatly enhanced. In addition, a long homologous arm designed in the homologous repair plasmid Phdr-TK (fLUC) further ensures accurate insertion of luciferase genes, and an off-target effect is avoided. The high consistency of the genetic background of the recombinant virus is ensured through a plaque purification technology. The method disclosed by the invention is convenient and quick, can be used for quickly constructing the herpes simplex virus (HSV) at lower cost, and is suitable for constructing and transforming various types of HSV viruses.

The invention discloses a Ralstonia solanacearum report filamentous bacteriophage, which is characterized in that a luciferase luxAB gene is inserted into a filamentous bacteriophage RSCq genome and then electrically transformed into Ralstonia solanacearum GMI1000 to construct the Ralstonia solanacearum report filamentous bacteriophage RSCqluxAB, and the gene sequence of the Ralstonia solanacearum report filamentous bacteriophage RSCqluxAB is as shown in SEQ ID The invention further discloses a construction method and application of the ralstonia solanacearum report filamentous phage. The Ralstonia solanacearum report filamentous bacteriophage has a detection rate of 85% for 42 Ralstonia solanacearum test strains, is suitable for detection of various Ralstonia solanacearum, only infects Ralstonia solanacearum, has strong specificity, fast detection speed and high detection sensitivity, and can make up for the defects of the existing Ralstonia solanacearum detection technology. Furthermore, the preparation and detection processes of the ralstonia solanacearum reporter filamentous bacteriophage are simple to operate, and the detection cost is low by using capraldehyde as a substrate.

A cell model for in vitro evaluation of compound-induced skin sensitization and a constructing method therefor. The method for constructing the cell model comprises the steps of: designing and constructing an sgRNA expression vector based on CRISPR / Cas9 vector system; designing and constructing a homologous recombinant vector capable of knocking a reporter gene linked to a self-cleaving peptide sequence into a specific site of the expression frame of the HMOX1 gene; co-transfecting the homologous recombinant vector, an hCas9 plasmid and the sgRNA expression vector into a cell, and carrying out monoclonal expansion to obtain the cell model. The present invention obtains a HaCaT cell model in which a luciferase gene is knocked in before the stop codon of the HMOX1 gene by combination of CRISPR / CAS9 and a monoclonal cell technique. The cell model realizes synchronous expression of the luciferase gene and the HMOX1 gene, thereby effectively distinguishing sensitizing compounds from non-sensitizing compounds.

This invention relates to an infectious clone of CA16 virus carrying the luciferase gene Nluc, its construction, and its applications. The P1 region of the CA16 genome from strain OP293089 was constructed into pcDNA, and the P2 and P3 regions were constructed into the pUC57 vector. The P1 and P2+P3 regions were recombined with the pSVA vector to construct an infectious CA16 clone. The Nluc luciferase reporter gene was then introduced to construct the pSVA-CA16-Nluc infectious clone. After rescuing the CA16-Nluc virus, its activity was experimentally verified to be similar to that of the parent virus. After continuous passage in Vero cells, the genetic stability of CA16-Nluc in in vitro cell culture was experimentally verified. Furthermore, this infectious clone can be used for high-throughput drug screening and has wide applications in basic research on viral replication and pathogenesis mechanisms, high-throughput drug screening, and neutralizing antibody screening.

Method for producing a spontaneous metastasis model is disclosed herein. The models, according to various embodiments herein, achieve a higher metastasis incidence rate, in an instance about 100% for mesenteric lymph node (mLN) invasion and about 80 to 90% for secondary organs. The models are kinetic model having a luciferase-based expression system that provide more statistically significant and robust data. The embodiments herein further include a method for producing the model for metastasis, comprising transplanting recombinant carcinoma cells having increased invasiveness and decreased tumorigenic properties and luciferase gene expression system, in one or more orthotopic positions in an animal.