459 results about "Reporter gene" patented technology

The invention provides an application of an AC2 gene and the like in improving the expression quantity of GFP (Green Fluorescent Protein) and / or inhibiting degradation of an mRNA (Messenger Ribonucleic Acid) attenuated substrate taking the GFP as a reporter gene. Researches find that AC2 has an mRNA decay inhibition function, can inhibit degradation of a nonsense-mediated mRNA decay substrate taking GFP as a reporter gene and degradation of an endogenous nonsense-mediated mRNA decay substrate of arabidopsis thaliana, can improve the expression quantity of mRNA of AT1G0106020, RPS6, SMG7, AT5G35490, AT5G64430, AT1G36730 and the like, can reduce the expression quantity of AT1G72450 and AT2G400, has interaction with Dcp1 and Dcp2, and can be used for inhibiting the expression quantity of mRNA of AT1G0106020, RPS6, SMG7, AT5G35490, AT5G6430, AT1G36730 and the like. The growth ability of the yeast in an auxotroph culture medium can be improved, the expression level of Dcp1 can be reduced, and a research result lays a foundation for clarification of AC2 functions.

The invention discloses a tomato LACS2 gene promoter and application thereof, and belongs to the technical field of biology. On one hand, the invention provides the tomato LACS2 gene promoter, and on the other hand, the invention provides application of the tomato LACS2 gene promoter in promoting target gene expression in plants. According to the invention, a tomato LACS2 gene promoter fragment (788bp) is constructed to drive an EGFP-GUS (Enhanced Green Fluorescent Protein-Glucuronidase) gene fusion expression vector. GUS histochemical staining and laser confocal microscopy observation prove that the promoter fragment can guide normal expression of the reporter gene in the tomato epidermis. Therefore, the LACS2 promoter can be used for researching development, quality improvement and postharvest preservation of tomato peels and specific expression of exogenous genes in the tomato peels.

The invention discloses a SbWRKY51 gene, a promoter and application of the SbWRKY51 gene and the promoter in improvement of salt tolerance of sorghum, and belongs to the technical field of plant genetic engineering. The SbWRKY51 gene is screened from sorghum, and the expression level of the SbWRKY51 gene is up-regulated after salt stress treatment. The protein coded by the gene belongs to a WRKY transcription factor family, has transcriptional activation activity, and can start the expression of a reporter gene. Sorghum overexpression strains and complementary strains of the gene are obtained, salt tolerance related physiological indexes are measured, overexpression of the gene can improve the seed germination rate, increase the seed root length, regulate ion balance and enhance the free radical scavenging capacity of plants, and the effect of the gene in the aspect of improving the salt tolerance is embodied. A further research finds that the SbWRKY51 gene can enhance the plant salt tolerance by regulating a lignin synthesis pathway. The technical scheme provides a basis for cultivating resistant plants, and has important guiding significance for enhancing the production potential of sorghum under a high-salt condition and promoting agricultural development.

The invention belongs to the technical field of genetic engineering, and particularly relates to an agrobacterium rhizogenes-mediated visualized genetic transformation method for cucurbitaceae plants, which comprises the step of transfecting the cucurbitaceae plants by using agrobacterium rhizogenes containing recombinant plasmids, and the recombinant plasmids are expression cassettes containing Ruby reporter genes. A visual agrobacterium rhizogenes-mediated transformation system of the cucurbitaceae plants is established by utilizing the RUBY gene, professional equipment or chemical treatment is not needed, red hairy roots can be visually screened to identify and induce the positive plants, and the method is simple, rapid and efficient and can be used for identifying and inducing the positive plants of the cucurbitaceae plants such as bottle gourds and the like. And the gene is transferred into an exogenous gene or is subjected to gene editing on the cucurbitaceae plant, so that the gene has important significance.

The invention discloses a cell and a method for detecting the activity of a GLP-1R / GIPR double-target agonist drug. The invention relates to a cell for co-expressing GLP-1R, GIPR and CRE-miniP-luciferase reporter genes, and provides a method for detecting the biological activity of a GLP-1R / GIPR double-target agonist drug based on the cell, and the method is simple to operate, short in test period, sensitive in reaction, small in variability, strong in specificity and high in accuracy, can be used for release inspection of products, and can be used for detecting the biological activity of the GLP-1R / GIPR double-target agonist drug. The method has important significance on quality control and clinical application of GLP-1R / GIPR double-target agonist drugs.

The invention belongs to the technical field of gene engineering, and particularly relates to a Cre reporting system based on single-domain antibody labeling and application of the Cre reporting system. According to the shark single-domain antibody with Cre specificity, when a DNA sequence of dLox-EGFP is provided, Cre can be marked and reported in real time, and reporter genes can be expressed only when Cre exists in cells, so that a strictly regulated gene expression platform is provided. The mark is completely reversible and is completely compatible with the currently used Cre / loxp system.

The invention discloses a D / Yama2019 genetic evolution pedigree influenza delta virus reverse genetic operating system. According to the invention, DNA fragments corresponding to segments of PB2, PB1, P3, HEF, NP, M and NS of a full-length genome of D / JY3002 are amplified by using seven pairs of designed primers, the DNA fragments are seamlessly cloned to a pHW2000 vector and a pCC1-DualPro vector to construct plasmids, the plasmids are co-transfected to a cell line, and the artificially rescued D / Yama2019 genetic evolution pedigree influenza delta virus rD / JY3002 is obtained; the genomic sequence of the rD / JY3002 is completely consistent with the genomic sequence of a naturally separated strain D / JY3002, the infection ability is equivalent, and the rD / JY3002 can be stably subcultured for at least five generations. According to the present invention, the constructed unidirectional expression plasmid pPolI-D / JY3002-PB1-240-GFP-240 is adopted to obtain the artificially rescued recombinant influenza delta virus rD / JY3002-GFP carrying the green fluorescence reporter gene, and after the host cell is infected, the GFP expression quantity is high and stable; the invention establishes an efficient and stable D / Yama2019 genetic evolution pedigree influenza delta virus reverse genetic operating system which can be used for developing an influenza delta virus vector presenting exogenous gene.

The invention provides a CsERECTA gene-based tea tree leaf shape regulation and control method and a tea tree light utilization rate improving method, which are characterized in that a sequence of a tea tree gene is cloned to obtain a complete sequence; then, the CsERECTA is subjected to bioinformatics analysis, expression of a GUS reporter gene is started by constructing a plant expression vector and connecting a target fragment, and a recombinant plant vector is used for genetic transformation of tobacco, so that the function of the CsERECTA is verified; the character of the CsERECTA transgenic line is analyzed, the leaf shape is determined, the method capable of improving the leaf shape of the tea tree is obtained, and a foundation is laid for cultivating a new variety and improving the light utilization rate of the tea tree by using the CsERECTA gene.

The invention relates to the technical field of molecular biology, in particular to a multichannel virus reporting system based on TAT trans-activation. The invention relates to a multichannel virus reporting system based on TAT trans-activation. The multichannel virus reporting system comprises an activated virus and a reporting cell, the activated virus is a human enterovirus A71 for expressing TAT protein; the reporter cells are cells transfected with reporter plasmids; the reporter plasmid comprises an LTR sequence and a plurality of reporter genes which are started to express by the LTR sequence. When the virus is activated to infect the reporter cell, the TAT protein carried by the virus enters the reporter cell to activate the expression of the downstream reporter gene, and the expression level of the reporter gene can reflect the virus replication condition, so that the screening or mechanism research of related antiviral drugs can be realized. According to the technical scheme, the technical problem that the flexibility, stability and high-throughput screening adaptability of a virus reporting system in the prior art are not ideal can be solved, and the virus reporting system has ideal popularization and application prospects.

The invention relates to the technical field of medicine screening, in particular to an active substance screening platform and application thereof. The active substance screening platform comprises a cell combination; the cell combination comprises immunoregulation screening cells and antioxidant screening cells; the immunoregulation screening cell is a mouse mononuclear macrophage RAW 264.7 carrying an NF-kappa B element conserved sequence and a reporter gene; the antioxidant screening cell is a porcine small intestine epithelial cell IPEC-J2 cell carrying an ARE element conserved sequence and a reporter gene. The invention provides a screening platform based on a specific cell combination, which can quickly obtain an active substance with immune regulation, antioxidation or two functions from a large number of candidate active substances, has the advantages of low cost and high efficiency, and has important application value.

The invention belongs to the technical field of biological medicine and molecular biology, and particularly relates to application of circ0005704 and related biological elements thereof in diagnosis and treatment of recurrent spontaneous abortion with unknown reasons. Researches find that differential expression and autophagy level of ULK1 in trophoblast cells of URSA patients are increased. High-throughput transcriptome sequencing is combined with bioinformatics analysis, and it is found that miR-26a-5p is down-regulated in URSA patients, is responsible for up-regulation of ULK1 and promotes autophagy of trophoblast cells, so that occurrence of URSA is increased. Through a high-throughput transcriptional set screening strategy and dual luciferase reporter gene analysis, it is found that circ0005704 enhances the expression of ULK1 through miR-26a-5p. In a word, the research of the invention shows that the circ0005704 / miR-26a-5p / ULK1 signal axis participates in the pathogenesis of URSA by adjusting the migration of the trophoblast cells, and a new target and scientific evidence are provided for the clinical treatment of URSA.

The present invention discloses an enterovirus infectious clone with a novel HiBiT reporter gene and a construction method and application thereof, including enterovirus EVA71, CVA16, CVA10 and CVA7 infectious clones with a novel HiBiT reporter gene, using HiBiT as a reporter gene for the first time on the basis of EVA71, CVA16, CVA7 and CVA10 infectious clones, preparing EVA71-HiBiT, CVA16-HiBiT, CVA7-HiBiT and CVA10-HiBiT infectious clones, further rescuing the reporter virus by the infectious clone, and verifying the biological activity and genetic stability of the reporter virus by cytopathic effect, plaque, WB and flow cytometry in vitro. The virus clone or reporter virus involved in the present invention has a wide range of application value in the field of anti-enterovirus drugs and high-throughput screening of neutralizing antibodies.

The invention specifically discloses a salix cheilophila in-situ genetic transformation method independent of a tissue culture system, and belongs to the field of plant genetic engineering. According to the method, semi-lignified branches of salix cheilophila are taken as receptors, vacuumizing treatment is performed by utilizing agrobacterium rhizogenes infection liquid, hairy roots are directly induced under a water culture condition after short-term dark culture, and positive identification is performed through reporter genes. According to the method, tedious sterile tissue culture operation is completely avoided, the transformation period is shortened to about 22 days, and the technical threshold and the time cost are remarkably reduced. By adopting the system, the positive rate can reach 54.33% when the pBI121 vector is used for transformation, and the efficiency is remarkably improved compared with that of an existing method. The method provides an efficient and reliable technical platform for root system gene function research, expression analysis and genetic improvement of salix cheilophila and related species thereof, and has high application potential.

The invention discloses an efficient streptomyces mobaraensis CRISPR / Cas9 editing method, and belongs to the technical field of gene engineering. According to the method, a CRISPR / Cas9 system in streptomyces mobaraensis is successfully constructed and optimized, a related gene ssti of aerial hypha growth delay is selected as a reporter gene, a CRISPR / Cas9 editing system is optimized, an optimal promoter P19570 of Cas9 and the most appropriate sgRNA of a targeted ssti gene are obtained, and on the basis, a repair template with the length of 2kb is selected, so that the highest single gene editing efficiency reaches 90.8%. Furthermore, an array containing two gRNA expression cassettes is constructed, simultaneous knockout of ssti and sapB genes in smY2019 is realized, and the knockout efficiency is 6.25%.

The present invention relates to a method for obtaining a genetically modified progeny of an avian animal by packaging a gene edited or genetically modified vector in a recombinant adenovirus and injecting into the dorsal aorta of an embryo when PGCs migrate to a developing gonad; compared with the existing disclosed method for transforming the PGCs in vitro, the operation method disclosed by the invention not only can stably and effectively infect the PGCs in vivo, but also has the advantages that more importantly, the probability of generating gene editing offspring is higher, and particularly, the breeding efficiency of cultivating the transgenic chicken offspring with KRT75L4 gene and MSTN gene knockout is higher; and on the other hand, the reporter gene or crispr-cas protein gene does not need to be integrated into the genome of the bird and the non-native protein does not need to be expressed, so that the operability is higher from the commercial level.

The invention relates to the technical field of gene engineering, in particular to a transgenic zebrafish system pTo12-efla-tjp1a-P2A-mCherry based on a Tol2 transposition subsystem and a construction method of the transgenic zebrafish system pTo12-efla-tjp1a-P2A-mCherry. The preparation method comprises the following steps: S1, constructing a Tol2 transposon expression vector pTol2-ef1a-tjp1a-P2A-mCherry which contains an ef1a promoter, a tjp1a gene, a P2A peptide sequence and an mCherry reporter gene; s2, carrying out in-vitro transcription to prepare Tol2 transposase mRNA; s3, mixing the expression vector with Tol2 transposase mRNA, and microinjecting the mixture into the single-cell stage embryo of the zebra fish; s4, performing fluorescence screening on the F0 generation embryos surviving after injection to obtain the Founder fish with positive transgenosis; s5, after the F0-generation positive fish is bred to be sexually mature, an F1 generation is obtained through mating, transgenic positive individuals with stable inheritance are screened out after identification, and a transgenic zebrafish strain is established. According to the transgenic zebrafish system pTo12-efla-tjp1a-P2A-mCherry based on the Tol2 transposon system and the construction method of the transgenic zebrafish system pTo12-efla-tjp1a-P2A-mCherry based on the Tol2 transposon system, efficient integration of exogenous genes is achieved by utilizing the Tol2 transposon system, and the transgenic zebrafish line capable of being stably inherited to the F1 generation is successfully obtained.

Provided herein are kits, compositions, systems, and methods for treating a subject with a tumor with kynurenine responsive bacteria (e.g., such that the bacteria reduce or eliminate said tumor). In certain embodiments, the kynurenine responsive bacteria comprise one or more nucleic acid sequences comprising: i) a first nucleotide sequence (e.g., not endogenous to said bacteria) encoding a kynurenine transcriptional regulator protein, and ii) a second nucleotide sequence (e.g., not endogenous to said bacteria or overexpressing an endogenous equivalent protein) encoding a kynurenine transporter protein. In particular embodiments, the kynurenine responsive bacteria further comprise at least one of the following: i) a third nucleotide sequence comprising a kynurenine responsive promoter upstream of a plasmid replication gene encoding a plasmid replication protein, ii) a fourth nucleotide sequence comprising a kynurenine responsive promoter upstream of a reporter gene encoding a reporter protein, and iii) a fifth or more nucleotide sequence comprising a kynurenine responsive promoter upstream of a gene essential for bacterial growth, or a gene codes a for therapeutic payload.

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 rubus cochinchinensis efficient genetic transformation method based on RUBY reporter gene and agrobacterium rhizogenes mediation, which realizes that the transformation period is shortened to be within 15 days and the transformation efficiency is improved to be more than 60% through direct explant treatment under a non-tissue culture condition. The method comprises the following specific steps: 1) preparing an explant material; (2) constructing an RUBY carrier and transforming agrobacterium rhizogenes; (3) infecting the explant by utilizing agrobacterium rhizogenes carrying a target gene; (4) co-culturing after infection and carrying out RUBY visual screening; 5) positive root molecular biology verification; (6) inducing positive roots into buds; according to the method, induction of the positive callus, the positive root and the positive bud of the rubus coreanus stem segment under the non-tissue culture condition is successfully achieved, a stable and efficient genetic transformation system is constructed, the method has the remarkable advantages of being easy and convenient to operate, low in cost, wide in applicability, high in reproducibility and the like, and a key technical support is provided for molecular breeding of the rubus coreanus.

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 discloses a method for detecting the biological activity of human immunoglobulin ADCP, which is characterized in that CHO-K1 / SPIKESARS2 is taken as a target cell, THP-1 cells (human leukemia monocytes which naturally express CD32a (Fc gamma RIIa) and CD64a (Fc gamma RIIa) and stably transfect NF-kB-Luc luciferase reporter genes) modified by genetic engineering are taken as effector cells, and a fluorescence reporter gene method is adopted for detection. Based on specific target cells and effector cells, the ADCP biological activity of two different human immune globulin preparations, namely intravenous injection human immune globulin and subcutaneous injection human immune globulin, can be accurately and reliably reflected by a drawn dose-effect curve through a detection system matched with specific human immune globulin solution concentration, and the method is simple and easy to implement. Cost is low, and practical application and popularization value is achieved.

The invention discloses a tissue-specific promoter screening vector and application thereof, and relates to the technical field of plant genetic engineering. The screening vector comprises an expression vector and a modified GUS (glucuronidase) reporter gene connected to the expression vector, the modified GUS reporter gene is Cat1muGUS, and the sequence of the Cat1muGUS is shown as SEQ ID NO. 3. According to the present invention, the Cat1muGUS variant is developed based on the Cat1GUS, the Cat1muGUS sensitivity is high, the problem of low dyeing screening efficiency caused by the fact that the Cat1GUS is not sheared in the dicotyledonous plant and the monocotyledonous special tissue can be overcome, in addition, the tissue-specific promoter screening vector is constructed based on the Cat1muGUS variant, and the screening efficiency is high; the method can be well applied to screening of all plant and all tissue specific promoters, and the screening efficiency is improved.

Provided herein are methods of sex sorting a plurality of insects based on sex-specific gene expression, the methods comprising (a) generating an exogenous nucleic acid molecule; (b) delivering the exogenous nucleic acid molecule into an insect body of the plurality of insects, wherein the exogenous nucleic acid molecule comprises a promoter region, a gender-specific splicing module, a reporter gene, and a transcription terminator; (c) detecting gender specific gene expression of the reporter gene; and (d) sorting the insects of the plurality of insects based on the detection of the gender-specific gene expression in step (c), thereby sex-sorting the insects based on the gender-specific gene expression.

There are SARS coronavirus 2 recombinant vectors derived from a GH clade SARS coronavirus 2 Korean isolate, which express distinct reporter genes, and the production method thereof. A full-length clone of a SARS coronavirus 2 Korean isolate or a derivative thereof, according to one embodiment, can be used as a standard material for evaluating the efficacy of therapeutic agents and vaccines in cell lines and animal models while maintaining infectivity and replication capacity when restored to viruses, can be used to develop a large-scale testing method for therapeutic agent development, and can be used to develop attenuated vaccine strains. In addition, a SARS coronavirus 2 recombinant vector derived from a Korean isolate or a derivative thereof, expressing a reporter gene, can be used for high-capacity, rapid drug screening in the development of antibody therapeutic agents and antiviral agents.

The invention discloses a bacillus constitutive strong promoter and application thereof, and relates to the technical field of biology, and the nucleotide sequence of the promoter is SEQ ID NO: 2. The mRNA transcriptional level of the reporter gene is improved; the method is used for starting a target gene to efficiently express amylase in bacillus subtilis. The method is also used for starting a target gene to efficiently express alkaline protease in bacillus licheniformis. The novel constitutive promoter with enhanced transcriptional activity is developed by taking P43 as a starting template, and under the same host and culture conditions, the absolute level of the constitutive promoter for driving expression of a reporter gene or a target product is higher than that of the current P43 promoter, so that core power is provided for performance improvement of a bacillus cell factory.

The invention discloses an SRCIN1 gene therapy vector for autism treatment and application of the SRCIN1 gene therapy vector, and relates to the field of autism treatment. The vector is a lentiviral vector and comprises an overexpression sequence of an SRCIN1 gene, the overexpression sequence is pSLentiti-CMV-Srcin1-P2A-EGFP-WPRE, and the titer of the overexpression sequence is 2.99 E + 08 TU / ml. The SRCIN1 gene therapy vector is delivered to the prefrontal cortex through brain stereotactic injection and is used for up-regulating SRCIN1 expression and improving social behaviors, cognitive functions and anxiety-like behaviors of autism model mice. The SRCIN1 gene therapy vector further comprises an EGFP (enhanced green fluorescent protein) reporter gene which is used for visualizing transfection efficiency. The vector disclosed by the invention is simple to prepare and high in targeting property, and can be used for remarkably improving the social ability and exploration ability of mice, improving infantile autism behaviors such as anxiety behaviors and repeated engraving behaviors of the mice and increasing PSD95 protein expression.

This invention relates to the field of plant genetic transformation technology, and provides a method for genetic transformation of crape myrtle stem segments with buds. The method includes: infecting crape myrtle explants with Agrobacterium tumefaciens containing the RUBY reporter gene, and co-culturing them in a co-culture medium; then transferring the co-cultured explants to a differentiation medium for differentiation culture; and inoculating the obtained clustered buds into a rooting medium to obtain genetically transformed positive seedlings; wherein the explants are crape myrtle tissue culture seedlings with stem segments containing buds. The crape myrtle stem segments used in this invention have a higher regeneration efficiency than leaves. The adventitious bud induction rate of crape myrtle leaves is 1.0%-4.7%, and the stem segment regeneration and proliferation coefficient is 5-20. A large number of positive plants can be obtained in the later stage, and no browning phenomenon occurs in any of the stem segments after infection.

The invention relates to a gene expression cassette and application thereof. The gene expression cassette comprises a tyrosinase gene, a transcriptional repressor protein gene, a melanin response promoter and a reporter gene, and the reporter gene is located at the downstream of the melanin response promoter and is regulated by the promoter. The gene expression cassette with a specific structure is designed, melanin in cells can be responded, a melanin biosensor can be further constructed by utilizing the gene expression cassette, the gene expression cassette has good specificity, and high-throughput screening of strains can be realized. Furthermore, ALE evolution is carried out on the strain for producing melanin, high-throughput screening is carried out based on a designed sensor, the strain for producing melanin with high yield is obtained, through shake-flask culture and 5L fermentation tank culture, the yield of melanin reaches up to 10.84 g / L and 29.04 g / L respectively, and an efficient and feasible solution is provided for large-scale production of melanin.

The present application relates to the field of genetic engineering, and in particular to a vector for analyzing plant promoter expression specificity, a preparation method and application thereof. The T-DNA region of the vector comprises, from 5' end to 3' end, a callus-specific promoter-driven selection marker gene expression cassette and a reporter gene expression cassette with a multiple cloning site; the callus-specific promoter-driven selection marker gene expression cassette comprises a callus-specific promoter, a selection marker gene coding sequence and a terminator. The vector of the present application utilizes a rice callus-specific promoter to drive the expression of a selection marker gene, can greatly reduce the non-specific interaction with the promoter of a target gene, improve the specificity of the expression of the target gene, and effectively reduce the biological safety risk caused by the selection marker gene in a transgenic plant, and has important application value in basic theoretical research and molecular breeding.

Provided herein are compositions and methods for real-time identification and isolation of base-edited cell populations. Also provided herein are methods for producing enriched isogenic lines of genetically modified cells, including base-edited human pluripotent stem cells. In particular, provided herein are methods utilizing transient expression of reporter proteins, the detectable signal of which is altered following base editing. Using the transient reporter with a base editor permits enrichment of isogenic populations of base-edited cells.