34 results about "Renilla luciferase" patented technology

The invention discloses a method for detecting promoter activity by utilizing double luciferase reporter genes. The method for detecting the promoter activity by utilizing the double luciferase reporter genes comprises step 1, building a pGL3-basic-MSTNpro recombinant which contains 7 sections of MSTN gene 5' control region fragments with different lengths; step 2, performing cultivation and planking on target cells, configuring a mixture of the pGL3-basic-MSTNpro of the step 1 and a lipidosome and enabling a renilla luciferase carrier pRL-TK to be served as an internal reference to perform cell co-transfection; step 3, performing detection on luciferase activity through the double luciferase reporter genes.

The invention discloses a novel multifunctional dual-luciferase reporter gene plasmid. The plasmid contains a luc firefly luciferase gene, a Rluc renilla luciferase gene, a SV40 poly(A) termination signal part and two independent multiple cloning sites, wherein the two independent multiple cloning sites are located on the upstream and the downstream of a luc firefly luciferase gene coder frame respectively. The dual-luciferase reporter gene plasmid containing detection gene and reference gene is successfully constructed, not only can the detection gene be transferred into a subject cell conveniently, but also the reference gene can be taken in correspondingly, the complexity of experiment operation and experiment errors are greatly reduced, the plasmid has the multiple cloning sites at thetwo ends of the firefly luciferase gene, the multiple cloning site located on the upstream of firefly luciferase can introduce a promoter sequence which can be used for genetic transcription adjustment, control and detection, and the multiple cloning site located on the downstream of the firefly luciferase can introduce a 3'UTR sequence which can be used for miRNA target identification, so that switching of different application of the same plasmid is achieved.

The invention discloses a method for detecting plasmids of polycyclic aromatic hydrocarbons in an environment and an application thereof, and relates to the fields of molecular biology and environmental biotechnology. According to the method provided by the invention, by extracting the promoter of a humanized AHR (Aromatic Hydrocarbon Receptor) gene, using a CV060 plasmid including a firefly luciferase and renilla luciferase reporter gene as a carrier and inserting the AHR gene promoter with a specific sequence, a recombined lentivirus plasmid of a reference internal type dual-luciferase reporter gene including the AHR gene promoter is built. The detection method provided by the invention is good in repeatability, high in sensitivity, reliable in detection result, and simple and convenient to operate, and can be widely applied to the detection of the polycyclic aromatic hydrocarbons in the environment.

The invention discloses a liposome medicinal composition with tumor targeting, in-vivo tracing and treating functions and a preparation method thereof. The medicinal composition is a liposome targeting medicament which is resistant to CD44 antibody coupling, has a molecular imaging function simultaneously, and can be used for monitoring the in-vivo distribution of a medicament in real time in a living body state. In particular, plasmids containing three fusion genes, including renilla luciferase, red fluorescent proteins and suicide gene thymidine kinase, are coupled to CD44 antibody mediation-resistant immunoliposome, the specificity of liposome nanoparticles in a liver cancer in-situ model of an in-vivo targeting NOD/SCID (Non-Obese Diabetic/Severe Combined Immune-Deficiency) mouse is monitored by detecting a renilla luciferase signal with a living body imaging system, and apoptosis of liver cancer cells is induced by applying target thymidine kinase of ganciclovir; and moreover, a targeted liposome can be coated with adriamycin for inducing apoptosis of the liver cancer cells. The liposome medicinal composition provided by the invention does not have any toxic or side effect, has small damage and a good effect, and is suitable to be applied for a long time.

The invention discloses a construction method of a human APP promoter-containing Luci cell line, and the specific method is as follows: construction of an expression vector containing a human APP promoter region-luciferase chimeric gene; cotransfection of constructed pGL3-APPP Luc-Puro plasmids and pRL-SV40 plasmids into a human embryonic kidney cell line HEK-293T cell line; laying and planting of transfected HEK-293T cells after trypsinization into a 10cm culture dish, puromycin pressurization screening to obtain five cell lines with genomes capable of stably integrating the human APP promoter region-luciferase chimeric gene, and detection of activity of firefly luciferase and renilla luciferase in the five cell lines to obtain the cell line containing the human APP promoter region-luciferase chimeric gene. The cell line prepared by the cell line construction method can be used for screening of new anti senile dementia compounds and therapeutic drugs for inhibiting the expression of human APP.

The invention discloses a dual-luciferase reporter gene plasmid applicable to screening of an orphan nuclear receptor activity regulating agent. The plasmid is a chimeric plasmid (p-TK(DR1)<3>-Fluc-hRluc) of DR1-containing firefly luciferase and the internal reference renilla luciferase. A construction method for the plasmid is to insert a fragment as shown in SEQ ID No. 1 into a hRluc-containingframework plasmid obtained after NheI/BglII double-digestion of the commercial plasmid pmirGLO. High-efficiency screening of the orphan nuclear receptor activity regulating agent can be realized through cotransfection of cells with the plasmid and an orphan nuclear receptor-overexpressed plasmid and activity detection via dual luciferases. The method is simple to operate, and the dual-luciferase reporter gene plasmid has high sensitivity and good specificity.

The invention relates to an establishment method of a renilla luciferase-carrying duck Tembusu reported virus and a product and application thereof. The establishment method comprises the following steps: restructuring an infectious cloned plasmid containing full-length cDNA of the duck Tembusu virus, inserting a gene of renilla luciferase into a site between 5'UTR and a structural gene of the TMUV virus to establish an infectious cloned plasmid, into which the RLuc gene is inserted, of the full-length TMUV reported virus, then obtaining a transcript of the reported virus by using a high-quality automatically-capped T7 in-vitro transcription kit, and transfecting BHK21 cells to rescue the reported virus. The renilla luciferase is excellent in sensitivity and convenient to use and can be quantitatively detected, so that the replication condition of the virus can be reflected by detecting the activity of the RLuc; and the renilla luciferase-carrying duck Tembusu reported virus can be applied to research in multiple aspects such as the life cycle and the replication mechanism of the TMUV, an anti-TMUV test, interaction between a viral protein and a host and other multi-aspect mechanisms.

The invention belongs to the field of biological technology, and in particular relates to an NF-kappaB dual-luciferase report cell line capable of stably expressing a pig derived TLR8 receptor gene, and a construction method of the NF-kappaB dual-luciferase report cell line. The cell line is an HEK293-NF-kappaB cell, in which a pTLR8 gene is transferred; and the HEK293-NF-kappaB cell is an HEK293cell, in which NF-kappaB luciferase and endodontic luciferase are introduced. The invention provides a stable cell line, which can be used for researching pTLR8 activated downstream NF-kappaB promoterreport gene detection. The invention also provides a biological material for an RNA-Seq technology to perform high-throughput screening on differential expression genes in the stable cell under the bTLR8 silent and activated states, and lays the foundation and a platform for further researching the species specificity of TLR8.

The invention relates to the technical field of biology, in particular to a verification method for detecting ath-miRNA170-3p targeted MSH2 by using a tobacco dual luciferase report system. The tobacco dual luciferase reporting system comprises fluorescein luciferase (F-Luc) and renilla luciferase (R-LuC). The R-Luc is used as an internal reference, and restriction enzyme cutting sites AvrII and AgeI are embedded into a 3 'UTR region of the FLuc. The method disclosed by the invention is beneficial to clarification of an arabidopsis thaliana MSH2 gene expression mechanism regulated and controlled by the athmiRNA170-3p and screening of the biomarker, and has important significance on research on response to adversity stress of the arabidopsis thaliana MSH2 gene regulated and controlled by the ath-miRNA170-3p.

The invention belongs to the technical field of biology and in particular relates to an NF-kB dual-luciferase reporter cell for stably expressing swine TLR5 receptor genes and a construction method thereof. A cell line refers to HEK293-NF-kB cells in which the swine TLR5 receptor genes are transferred, and the HEK293-NF-kB cells refer to HEK293 cells in which NF-kB firefly luciferase and renilla luciferase are transferred. The invention provides the stable cell line used in detection for researching pTLR5 activated downstream NF-kB promoter report genes. The invention also provides a biological material for performing high-throughput screening on differentially expressed genes in stable cells in quiesced and activated states in an RNA-Seq technology, and lays a foundation and platform forfurther researching species specificity of TLR5.

A kit is used for diagnosing NPM1 antibodies, ranilla luciferase genes and NPM1 antigen genes are recombined into fusion vectors, and through a cell transformation technology, the fusion vectors are guided into a mammalian cell system to conduct luciferase-NPM1 antigen fusion protein expression; gene recombination cells are disrupted, luciferase-NPM1 antigen fusion proteins are obtained, NPM1 antigens are coated with a 96 elisa plate, and to-be-detected serum is added; the antibodies in the serum are combined with the antigens on the plate to be adsorbed to the plate; and the luciferase-NPM1 antigen fusion proteins are added, through an immunoprecipitation reaction, the luciferase-NPM1 antigen fusion proteins are combined with the NPM1 antibodies in the serum of a patient and are adsorbedto the plate, and finally a luciferase substrate is added to detect the fluorescence intensity. According to the fluorescence intensity, the content of the NPM1 antibodies can be reflected to providereference for diagnosis of prostatic cancer; and a method is quick, simple and convenient and has high sensitivity.

The invention discloses a construction method for stably expressing a Zika virus replicon containing renilla luciferase Rluc and an application thereof. According to the invention, a replication subsystem of the Zika virus containing the renilla luciferase Rluc is constructed, so that the replication subsystem can be stably expressed in cells, and the replication condition of the virus can be detected by detecting a renilla luciferase Rluc signal. Meanwhile, the method can be used for antiviral drug screening.

The invention discloses a high-throughput drug screening model using a human pregnane X receptor (hPXR)-mediated dual-luciferase reporter gene technique. The invention discloses a method for screening a human pregnane X receptor stimulant and an antagonist; by virtue of the method which is a new drug screening method, active drug ligands having regulating effect on a target gene, namely cytochrome CYP450 gene, of the PXR. A human embryonic renal HEK293T cell is subjected to the instantaneous co-transfection of an hPXR expression plasmid, a firefly luciferase reporter gene plasmid and an internal reference renilla luciferase plasmid by virtue of the method. The activities of the firefly luciferase and the renilla luciferase are simultaneously detected to show the regulating effect of the drug on the PXR activity, so as to establish a method for screening the hPXR stimulant and the hPXR antagonist. On one hand, the method can screen out compounds having an hPXR stimulating activity at high throughput from a massive compound library so as to avoid a risk that adverse drugs interact caused by the combined use of new drugs; and on the other hand, the method can screen out compounds having PXR antagonistic activity to inhibit the inducing effect of ligands on drug metabolic enzyme and transporter, so as to reduce the occurrence of adverse DDI and to reverse the drug resistance of anticancer drugs.

The invention discloses a PPAR gamma receptor stimulant as well as a preparation method and application thereof. One receptor stimulant is ursolic acid extracted from asiatic cornelian cherry fruit. The preparation method specifically comprises the following steps: step 1, preparing an asiatic cornelian cherry fruit extract; and step 2, separating and purifying the asiatic cornelian cherry fruit extract obtained in the step 1 to obtain the receptor stimulant. The invention provides a novel natural PPAR gamma receptor stimulant, and the relative average intensity of the firefly luciferase activity and the internal control renilla luciferase activity of the compound is equivalent to that of the full stimulant rosiglitazone of the PPAR gamma receptor under the concentration of 3 microns; theresearch result provides a lead compound for research and development of novel natural antidiabetic drugs.

The invention discloses a recombinant plasmid containing an HDAC1 (histone deacetylase 1) gene promoter and a reporter gene as well as a construction method and the application of the recombinant plasmid. A promoter specific sequence of a histone deacetylase 1 gene is cloned and recombined with the promoter specific sequence to a firefly luciferase reporter gene vector so as to construct a recombinant plasmid vector of the HDAC1 reporter gene; the constructed vector and a reporter gene vector of renilla luciferase are co-transfected into human prostate cancer cells PC3, and the transcriptional activity of an active reaction HDAC1 gene promoter of firefly and renilla luciferase is detected at the same time, so that a method for targeted screening of tumor inhibiting drugs is established. The method has excellent application prospects in the aspects of screening tumor inhibiting drugs, evaluating the possible influence of to-be-detected drugs on tumors, researching the action mechanism of the drugs and the like.

The invention belongs to the technical field of biology and specifically relates to an NF-kappa B dual-luciferase reporter cell system capable of stably expressing pig-origin RIG-I receptor genes anda construction method thereof. The cell system is an HEK293-NF-kappa B cell which the pig-origin RIG-I (pRIG-I) receptor genes are transferred into, and the HEK293-NF-kappa B cell is an HEK293 cell which NR-kappa B firefly luciferase and internal reference renilla luciferase are guided into. The stable cell system disclosed by the invention can be applied to researching detection on downstream NF-kappa B promoter reporter genes activated by pRIG-I. The invention further provides a biological material for an RNA-Seq technology to screen stable cell internal differential expression genes under silent and active states of the pRIG-I in high throughput; furthermore, a foundation and a platform are established for further researching species specificity of the pRIG-I.