585 results about "Reporter gene" patented technology

The invention belongs to the field of biotechnology and medicine, and discloses mRNA (messenger Ribonucleic Acid) for coding luciferase and application thereof. The mRNA for coding the luciferase comprises an ORF sequence for coding the luciferase, and the ORF sequence for coding the luciferase is a nucleotide sequence as shown in one of SEQ ID NO: 10 to SEQ ID NO: 13. The invention provides a brand-new mRNA for coding luciferase, and the mRNA is stable in structure, not easy to degrade, high in translation efficiency, relatively high in protein yield, relatively high in biological availability and safety by reasonably optimizing a nucleic acid sequence of a functional region, can be applied as a reporter gene, and has good potential.

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 provides a construction method and application of a high-efficiency expression system based on a Seali forest virus replicon. The method comprises the following steps: constructing a saRNA system for expressing a reporter gene; constructing to obtain replicase providing plasmids in the taRNA system; constructing to obtain an antigen providing plasmid in the taRNA system; the method comprises the following steps: respectively linearizing pSFV-CMV-T7-X, pSFV-CMV-T7-TR-X and pSFV-CMV-T7-replicase, and then carrying out in vitro transcription to prepare mRNA (Messenger Ribonucleic Acid); transfecting the prepared mRNA into a host eukaryotic cell for expression according to a certain proportion to obtain an antigen protein; and detecting the antigen protein to evaluate the expression efficiency of the saRNA and taRNA system based on the SFV replicon. According to the self-amplification DNA and RNA vaccine based on the virus replicon, the development period can be shortened, and the vaccine antigen can be expressed at a high level.

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 present invention discloses a method for selective crop management in real time. The method comprises steps of: (a) producing a biosensor plant, said biosensor plant comprises a visual biomarker, said biomarker is encoded by at least one modified genetic locus comprising (i) preselected reporter gene allele having a phenotype detectable by a sensor, and (ii) a regulatory region of a preselected gene allele responsive to at least one parameter or condition of said plant or its environment, said regulatory region is operably linked to said reporter gene, such that the expression of said reporter gene phenotype is correlated with the status of said at least one parameter or condition of said biosensor plant or its environment; (b) acquiring image data of a target area comprising a plurality of said biosensor plants via said sensor and processing said data to generate a signal indicative of the phenotypic expression of said reporter gene allele of said biosensor plant; and (c) communicating said signal to an execution unit communicably linked to the sensor, said execution unit is capable of exerting in real time a selective monitoring and / or treatment of said target area or a portion thereof comprising said biosensor plants, said treatment is being responsive to said status of said parameter or condition of the biosensor plant or its environment. The present invention further discloses systems and plants related to the aforementioned method.

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 relates to the technical field of plant genetic engineering, in particular to a genetic transformation method of prunus mume line plum blossom leaf calluses. The method comprises the following steps: firstly preparing a plum blossom genetic transformation receptor, then transforming agrobacterium tumefaciens by taking an RUBY gene as a reporter gene to prepare an infection bacterium solution, then carrying out callus infection and co-culture, and finally screening to obtain the resistant callus. According to the method provided by the invention, the resistant callus can be identified in an observation or PCR (Polymerase Chain Reaction) detection mode. The plum blossom callus is successfully infected, the red callus expressed by the RUBY reporter gene is obtained, the inductivity is high, and a good basis is provided for establishment and genetic improvement of a plum blossom genetic system.

The invention provides application of a P8 gene, or a P8 gene coding protein, or a recombinant vector or host bacteria containing a P8 gene coding region in improvement of GFP (Green Fluorescent Protein) expression quantity and / or inhibition of degradation of an mRNA (Messenger Ribonucleic Acid) attenuated substrate taking GFP as a reporter gene. The research finds that the P8 has a silence 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 (NMD) substrate of arabidopsis thaliana, can improve the mRNA expression quantity of AT1G0106020, RPS6, SMG7, AT5G35490, AT5G64430, AT1G36730, AT4G3900 and AT5G22570, and can reduce the expression quantity of AT1G72450 and AT2G400, and the research result lays a foundation for clarification of the function of the P8.

The invention discloses a Myo6 gene enhancer and application thereof, and belongs to the field of gene engineering. The Myo6 gene enhancer can drive the expression capacity of a reporter gene EGFP or Myo6 gene in the inner ear to be obviously enhanced, can be used for biological materials such as recombinant DNA, vectors or adenoviruses and the like, promotes the expression of the Myo6 gene or other genes in the inner ear so as to carry out gene expression regulation and control research, and is expected to be used for gene therapy of deafness.

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 provides a genetic transformation method suitable for shiny-leaved yellowhorn hairy root induction. The genetic transformation method comprises the following steps: culturing shiny-leaved yellowhorn seedlings infected by agrobacterium rhizogenes, preparing agrobacterium rhizogenes infection liquid, infecting the agrobacterium rhizogenes, inducing transgenic hairy roots by the agrobacterium rhizogenes and culturing. According to the method, a receptor material, a red tag gene RUBY and a green fluorescent gene GFP which are rapidly cultured and are suitable for agrobacterium rhizogenes infection are used as reporter genes, induction and transformation of hairy roots are carried out through agrobacterium rhizogenes mediation by utilizing a non-tissue culture infection and induction culture method, and the hairy roots are obtained. The cultivation period and cost of traditional plant hairy root induction are reduced, and a hairy root genetic transformation method suitable for shiny-leaved yellowhorn is established.

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 relates to the technical field of molecular biology, and particularly discloses application of miR-615 in ovarian granular cells. The miR-615 is applied to any one of the following items: preparing a product for promoting the activity and / or proliferation of ovarian granular cells; preparing a product for promoting generation of estradiol and / or progesterone; preparing a product for promoting the expression of the 3 beta-HSD and / or StAR gene; preparing a product for inhibiting ACSL4 gene expression; and preparing a product competitively combined with the lnc-GAM. According to the invention, porcine ovarian granular cells are used as a material, the expression level of lnc-GAM-miR-615 / ACSL4 regulatory axis is up-regulated or down-regulated in the porcine ovarian granular cells by using gene overexpression and interference technologies, and the expression of the lnc-GAM-miR-615 / ACSL4 regulatory axis is determined by detecting cell proliferation activity, hormone secretion level and apoptosis gene expression in combination with a dual-luciferase reporter gene test. And the regulation effect of lnc-GAM mediation on the apoptosis of the porcine ovarian granular cells is researched.

Cas-cleavable reporter systems, CRISPR-Cas systems thereof, and methods of use thereof in CRISPR-Cas based diagnostics. Cas-cleavable reporter systems may generate a luminescent, fluorescent, or other detectable signal upon Cas-collateral cleavage of one or more reporter system components. CRISPR-Cas systems may comprise a Cas protein having collateral cleavage activity, guide molecules, and the Cas-cleavable reporter system. Cas-cleavable reporter systems may be a split luciferase reporter system, at least one element of which is bound to beads. For multiplexing, CRISPR-Cas systems may comprise a Cas-cleavable quenched reporter system, optionally a Cas-cleavable quenched fluorescent reporter system, a Cas protein having collateral cleavage activity, and target-specific guide molecules attached to color-coded beads. CRISPR-Cas systems may further comprise amplification reagents. Methods may apply said CRISPR-Cas systems to flow cell, well-plate, or other devices, and may measure detectable signals by microscopic, plate reader, or other methods.

The present invention discloses an Ush1c gene enhancer and its application, belonging to the field of genetic engineering, and the nucleotide sequence of the enhancer is shown in SEQ ID No. 1. The mouse embryo microinjection system is used to confirm that the sequence is an enhancer, which can significantly enhance the transcription activity of the EGFP protein gene in the inner ear. Compared with other sequences of similar size, the enhancer sequence fragment can drive the reporter gene or Ush1c to express significantly in the inner ear, and specifically express it in inner and outer hair cells, and is suitable for recombinant DNA, vectors or adenovirus and other biological materials, and is used to promote the transcription of EGFP gene or other genes in the inner ear.

The invention discloses a jujube genetic transformation method using anther to induce callus as a receptor. The method comprises the following steps: culturing sterile anther to obtain callus; constructing a plant recombinant overexpression vector containing a target gene, and transferring into agrobacterium to obtain agrobacterium dip dyeing liquid; the callus is dip-dyed with an agrobacterium dip-dyeing solution, and then the dip-dyed callus is co-cultured under a dark condition; and then putting the callus on a culture medium containing hygromycin for screening, and screening and culturing for 20-30 days to obtain the homozygous resistant callus. According to the method, the callus induced by the Jing 39 jujube anther is used as a receptor for the first time, the method of the Jing 39 jujube genetic transformation system is established, the transformation frequency is high, meanwhile, screening can be assisted by means of a 35S:: EGFP fluorescent reporter gene marker, and convenience and rapidness are achieved. The method provides convenience for jujube gene function analysis and jujube genetic improvement.

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 invention discloses a dual-luciferase reporter gene detection system and application thereof. The detection system comprises Gaussian luciferase, Pleuroma xiphias luciferase and a substrate, and the substrate is coelenteran or a derivative of coelenteran. Compared with other dual-luciferase reporter gene detection systems, the dual-luciferase reporter gene detection system has stronger signals and simple reaction conditions.

The invention discloses a reverse genetic operating system of fowl adenovirus type 4 and application of the reverse genetic operating system. Belongs to the technical field of virus genetic manipulation. The FAdV-4 full-length genome is amplified in a segmented mode through a PCR method, segmented amplification products are cloned to pOK-CMV vectors through the homologous recombination technology, four library plasmids covering the whole gene are successfully obtained, and LMH cells are transfected to successfully rescue the virus. Meanwhile, the reporter gene NanoLuc is inserted into the FAdV-4 genome, so that the recombinant FAdV-4 for expressing the NanoLuc is successfully rescued. The invention provides an efficient technical platform for deep development of foundation and application research of FAdV-4, and has important scientific application value.

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 relates to the technical field of plant genetic engineering, and particularly discloses a construction method of a VIGS silencing system for agastache rugosus gene function research, which comprises the following steps: constructing a TRV2-PDS or TRV2-CHLH visual vector by taking indicator genes PDS and CHLH for chlorophyll synthesis as reporter genes; the method comprises the following steps: transforming a TRV2-PDS or TRV2-CHLH visual vector into agrobacterium, and performing injection infection on cotyledons of a target plant to construct a VIGS silencing system. According to the invention, cloning, amplification and vector construction are carried out on the isopuleone isomerase gene (ArIPI gene) in the agastache rugosus, and the function of the ArIPI gene is verified in a plant body, so that a scientific basis is provided for improving and cultivating the agastache rugosus and constructing genetic engineering.

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 invention provides a multiple nucleic acid detection technology based on CRISPR / Gs12 system mediation and application, different Gs12 nuclease are respectively combined with corresponding specific crRNA and single-stranded DNA reporter genes, and then a recombinase polymerase amplification (RPA) method is combined, so that quadruple, triple or double specific accurate detection of target nucleic acid molecules can be realized. Therefore, a nucleic acid detection technology and a kit capable of simultaneously distinguishing the Seneca virus (SVA), the hog cholera virus (CSFV) and the African hog cholera virus (ASFV) are developed. The method provided by the invention can realize rapid multiple detection of target nucleic acid, and has good popularization and application values in the field of nucleic acid detection.

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.