157 results about "CYP2C19" patented technology

The invention relates to a medicine metabolic relevant loci detection method, which comprises the following steps: extracting genome DNA from human samples; respectively designing a Taqman probe pair and a primer pair according to at least two medicine metabolic relevant genes; respectively marking the 5' end and the 3'end of the Taqman probe pair with fluorescence reporting genes and fluorescence quenching genes; carrying out fluorescence quantitative PCR augmentation on the genome DNA; and judging whether the medicine metabolic relevant genes have the mutation according to the fluorescence quantitative PCR augmentation results. Preferably, the number of the medicine metabolic relevant genes is four, the Taqman probe pair and the primer pair are used for detecting a loci rs1057910 of a gene CYP2C9, a loci rs4244285 of a gene CYP2C19, a loci rs4986893of a gene CYP2C19, a loci rs1065852 of a gene CYP2D6 and a loci rs28371759 of a gene CYP3A4. The invention has the advantages of ingenious design, simple operation and accurate and reliable detection results, and provides the reference frame for determining whether professional doctors are needed to be consulted so as to make sure the medicine can be taken or not or the proper dosage and the like when a certain medicine is taken.

The invention relates to a kit and a method for detecting polymorphism of CYP2C19 gene, and belongs to the field of fluorogenic quantitive PCR (Polymerase Chain Reaction). The kit comprises a detection primer and a fluorescent probe, which comprise at least one group of specific primer and specific Taqman fluorescent probe of CYP2C19 gene CYP2C19*2 polymorphism, specific primer and specific Taqman fluorescent probe of CYP2C19 gene CYP2C19*3 polymorphism, and specific primer and specific Taqman fluorescent probe of CYP2C19 gene CYP2C19*17 polymorphism. The kit is adopted for detecting the CYP2C19 gene, the sensitivity and specificity are both obviously improved, the detection time is short and the predication of medicament dosage is facilitated.

The invention discloses a CYP2C19 and ABCB1 gene SNP detection liquid-phase chip and a detection method thereof. The liquid-phase chip comprises microspheres and amplification primer, wherein the microspheres are coated with specific anti-tag sequences respectively; and the anti-tag sequences are selected from sequences from SEQ ID NO.7 to SEQ ID NO.12. As for three pairs of ASPE primers formed by specific sequences of SNP sites of a target gene and tag sequences of 5' ends, the specific sequences are SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3, SEQ ID NO.4, SEQ ID NO.5 and SEQ ID NO.6 respectively. The fluorescent signal value detected by the liquid-phase chip is greatly improved, so that the detection sensitivity is further improved, the signal-to-noise ratio is strengthened, and the detection result is more accurate and reliable.

The invention belongs to the gene detecting technology in clinical detecting technologies of the bio-medical field and particularly relates to a kit for quickly detecting polymorphism of a human CYP2C19 gene and a using method of the kit. The kit comprises three pairs of primers, three probes, six PNA (pentose nucleic acid) sequences, a pair of beta-Actin interior label primers and an interior label probe, and further comprises a PCR (polymerase chain reaction) buffer solution of Mg<2+>, a dNTP mixture, Taq enzyme and ddH2O. The kit can be utilized to carry out qualitative detection on a polymorphic site of the CYP2C19 gene and carry out two-tube PNA-PCR fluorescent amplified reaction according to the SNP site and can carry out result judgment according to a condition whether a two-tube fluorescence curve is started or not without producing personal errors, so that a false positive rate and a false negative rate are low. And moreover, the kit can be used to realize high-flux detection easily and is capable of clinical use.

The invention relates to a preparation method, a detection method and application of a probe drug composition for determination of metabolic activity of cytochrome P450. The composition mainly comprises a preparation made with a specific probe with major isoforms of CYP450, i.e. CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP2E1 and CYP3A4, as an active component. Cocktail probe drug solution is prepared, the probe drug composition is injected into an animal or liver microsomes for in vitro co-incubation, and the concentration of each probe drug is determined to assess the metabolic activity of the CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP2E1 and CYP3A4. In the early stage of research and development of new drugs, the effects of the drugs on the activity of each isoform of the cytochrome P450 are screened in a high-throughput way, and the interactions of the drugs can be predicted. In the stage of clinical research, the testing can be performed with the probe drug composition in an in-vivo probe method, and the effects of the drugs on the in-vivo metabolic activity of different isoforms of the human liver CYP450 can be examined.

The invention belongs to the field of biotechnologies and medicine, and provides a specific primer used for human CYP2C19 gene polymorphism detection. The nucleotide sequences of the primer are shown in SEQ ID NO:1-9. The invention further provides a kit for human CYP2C19 gene polymorphism detection. The primer and the kit are used for human CYP2C19 gene polymorphism detection and have the advantages of being high in specificity and sensitivity, easy and fast to operate and safe and having high flux, and result interpretation is objective.

The present invention discloses a gene detection kit using the multiplex PCR technology combined with the SNP sensitive molecular switch technology for genotyping of cytochrome 4502C19 gene, an amplification method and a detection method. Three common polymorphic sites on the CYP2C19 gene significantly changing the product activity are genotyped by the kit, wherein the three common polymorphic sites include: rs4244285SNP site, rs4986893 site and rs12248560 site. The kit includes a wild-type 2*PCR buffer, a 2*mutant amplification buffer, and a polymerase. The two buffers respectively includes a sequence-specific primer and an internal reference primer corresponding to SNP wild and mutant phenotype, and can complete genotyping for the three SNP sites in two multiplex PCR reactions, and thus the CYP2C19 gene is genotyped, and molecular biological evidence is provided for the gene-expressed enzyme activity prediction.

The invention belongs to the technical field of biology, and particularly relates to a kit for detecting efficient typing of gene loci related to clopidogrel drug resistance reaction. The kit is based on a Sequenom MassARRAY typing system, and comprises PCR (polymerase chain reaction) amplification primers and single-base extension primers of gene loci CYP2C19*2 (rs4244285) and CYP2C19*3 (rs4986893). The sequences of the PCR amplification primers of gene loci are respectively disclosed as SEQ ID NO.1, SEQ ID NO.2, SEQ ID NO.3 and SEQ ID NO.4. The sequences of the single-base extension primers of gene loci are respectively disclosed as SEQ ID NO.5 and SEQ ID NO.6. Before the subject takes clopidogrel, whether the gene loci of the subject are mutated is detected to instruct the individualized reasonable application of the subject, thereby providing references for individualized antiplatelet therapy.

The invention provides a sequencing primer for qualitative detection of cytochrome oxidase CYP2C19 genetic typing and a kit of the sequencing primer and belongs to the field of external nucleic acid detection. The kit comprises uracil DNA (Deoxyribonucleic Acid) glycosylase, Taq polymerase, PCR (Polymerase Chain Reaction) reaction liquor, PCR amplification primers, pyrophosphoric acid sequencing primers and positive control products. The kit is high in sensitivity and good in specificity, PCR products can be simply treated for a pyrophosphoric acid sequenator for sequencing, operation is simple, reaction time is short, the sensitivity of the kit is higher than that of a golden standard-capillary electrophoresis sequencing, and the kit is suitable for use for mutation analysis.

The invention provides a novel method for detection of human CYP2C19 gene polymorphism. The method comprises the following steps: (1) an ARMS primer is designed. The 3' end of the ARMS primer is a mutation site, and any missense in first-third nucleotide positioned upstream of a mutation site is mutated; (2) after allelic ARMS primer is designed for each SNP, an artificial nucleotide sequence with no homology with human gene sequence is respectively introduced into the 5' end of each of the two specific ARMS primers; (3) according to the introduced general artificial sequence, a Taqman probe which is completely complementary and paired up with the intermediate sequence fragment of the general sequence is designed, and a specific fragment amplified by the ARMS primer is specifically identified. By the above detection method and the kit, the capability of the allelic specific primer in distinguishing different alleles can be greatly enhanced; and accuracy of detection results is guaranteed, cost is greatly saved and efficiency is raised.

The invention provides a detection probe for an SNP (Single Nucleotide Polymorphism) of a human CYP2C19 gene. A detection primer consists of a specific upstream and downstream primer pair of the gene and a specific Taqman double fluorescent probe which are respectively used for correspondingly detecting SNPs at CYP2C19*2, CYP2C19*3 and CYP2C19*17 sites. The detection probe has the characteristics of quick detection and high specificity; furthermore, the detection method is simple; positive contrast and negative contrast which are necessary to conventional fluorescent quantitation PCR are eliminated, so that the operation steps and the experimental cost are reduced; the subsequent clinical treatment strategy can be guided more quickly and better.

The invention relates to the field of molecular biology, and discloses a method for detecting gene mutation of human cytochrome P450 CYP2C19, which comprises the following steps of: performing amplification on a sample genome by using primers, wherein the primers comprise upstream and downstream primers for detecting M1 mutation, and upstream and downstream primers for detecting M2 mutation, the nucleotides of the upstream and downstream primers for detecting M1 mutation are shown as SEQ ID No.1 and SEQ ID No.2, and the nucleotides of the upstream and downstream primers for detecting M2 mutation are shown as SEQ ID No.5 and SEQ ID No.6; and setting a reference for quality control, and sequencing an amplification product by using sequencing primers of which the nucleotides are shown as SEQ ID No.9 and SEQ ID No.10. The invention also provides a kit for detecting gene mutation of human cytochrome P450 CYP2C19. According to detection results of the method, guidance and adjustment of clinical medication schemes are carried out, a basis is provided for clinical individualized medication, the treatment effect can be improved and the toxic and side effect risks can be reduced.

The present invention relates to a method for predicting the resistance of a human subject to clopidogrel, which comprises detecting the presence or absence of a A allele at position 636 of exon 4 in the CYP2C19 gene, wherein the presence of the A allele is indicative of a clopidogrel resistance. The present method may be very useful in predicting the resistance of a human subject to clopidogrel and contribute to more effective chemotherapy for patients having coronary artery disease and drug-eluting stent.

The invention discloses a CYP2C19, CYP2C9 and VKORC1 genotyping multiplex amplification system and detection kit. The amplification system comprises two forward primers and a reverse fluorescent primer of six SNPs (single nucleotide polymorphisms) respectively and can simultaneously amplify the six SNPs. The system is characterized by achieving one-tube amplification of three SNPs on the gene CYP2C19, two SNPs on the gene CYP2C9 and one SNP on the gene VKORC1 through multiple PCR (polymerase chain reaction). In particular, the amplification system can achieve direct amplification of blood and blood spot samples and dispenses with the step of extracting DNA (deoxyribonucleic acid). The amplification system can integrate the UDG-dUTP antipollution measure and can effectively prevent product pollution. The detection system is comprehensive in site detection, is simple and convenient to operate, has high specificity, high sensitivity and strong reliability, is low in cost and has the capacity of mass detection.

The invention belongs to the technical field of gene engineering and discloses a composition of primers and probes for detecting polymorphism of human CYP2C19 gene, a kit containing the composition of the primers and the probes, and a fluorescence PCR method for detecting the polymorphism of the human CYP2C19 gene by using the composition of the primers and the probes or the kit. Based on a TaqMan fluorescence PCR technology, the primers, the probes and the kit are simple and rapid, and are high in sensitivity; in addition, through reasonable collocation of the primers and the probes, the interaction between the primers, the interaction between the primers and the probes and the interaction between the probes can be effectively avoided; the detection errors are reduced. When the primers, the probes and the kit, provided by the invention, are used for detecting the polymorphism of the human CYP2C19 gene, the primers, the probes and the kit has the advantages that the sensitivity is high, the specificity is high, the operation is simple, rapid and safe, the result is simply and intuitively determined and read, and blood samples or dried blood spots samples on filter paper can be directly used as templates.

The invention relates to a kit and a method for detecting CYP3A5 gene polymorphism, belonging to the field of fluorescent quantitative PCR (Polymerase Chain Reaction). The kit comprises primers for detection and fluorescent probes for detection, wherein the primers for detection and the fluorescent probes for detection comprise at least one group of specific upstream and downstream primers for detecting CYP3A5*3 polymorphism of the CYP3A5 gene as well as a Taqman fluorescent probe, specific upstream and downstream primers for detecting CYP2C19*4 polymorphism of the CYP3A5 gene as well as a Taqman fluorescent probe, specific upstream and downstream primers for detecting CYP2C19*6 polymorphism of the CYP3A5 gene as well as a Taqman fluorescent probe, and specific upstream and downstream primers for detecting CYP3A5*7 polymorphism of the CYP3A5 gene as well as a Taqman fluorescent probe. The detection sensitivity and specificity of the kit are remarkably improved, the detection time is short, and prediction of drug dose and curative effect is facilitated.

The invention provides a primer combination for detecting gene polymorphic sites relevant to clopidogrel resistance, wherein the sites comprise the polymorphic sites including CYP2C19*2, *3, *4, *5, *17 and the polymorphic site 3435C>T of ABCB1. According to the scheme, multiple PCR primers are utilized for amplifying gene segments where relevant gene loci are located, after the amplification product is treated, single base extension is carried out on the to-be-detected loci, then molecular weight difference detection is carried out on the extension product by adopting a flight time mass spectrum, through data analysis, the result of the relevant gene for the clopidogrel resistance of a patient is rapidly detected. The invention further provides a detection product prepared by adopting the primer combination and applications of the detection product, and thus references are provided for the individualized medication of clopidogrel.

The invention discloses a kit for simultaneously detecting multisite mutation of genes CYP2C19 and CYP2D6. A group of Taqman allele resolution analysis method-based specific primers and probes is obtained by meticulous designing, multiple verification, screening and optimization, and 8 functional variations of 4 types, i.e. single base displacement, deletion mutation, gene deletion and duplication mutation, of genes CYP2C19 and CYP2D6 can be detected; from DNA extraction to fluorescent PCR and then to result acquisition, 4 hours is required only, and the manual operation time is shorter than 2 hours. The kit containing the primers has the advantages of time saving, convenience, high sensitivity, capability of ensuring that the positive coincidence rate and negative coincidence rate of a sample are both over 99 percent, and the like.

The invention relates to the technical field of DNA detection and provides an SNP typing method and kit. A to-be-tested SNP site comprises SNP sites in CYP2C9, VKORC1 and CYP2C19 genes. The SNP typing method comprises the following steps: replacing a fluorescently-labeled oligonucleotide probe with a non-fluorescently-labeled oligonucleotide probe, performing one-time connection sequencing reaction on to-be-sequenced library molecules and then obtaining sequence information of the to-be-tested SNP site through connection sequencing. The SNP typing kit comprises a CYP2C9 specific primer pair, a VKORC1 specific primer pair, a CYP2C19 specific primer pair and the non-fluorescently-labeled oligonucleotide probe. The SNP typing method and kit provided by the invention have the benefit that through sealing non-specific binding sites in the to-be-sequenced library molecules, a false signal in a result obtained by the connection sequencing is reduced, so that the accuracy of SNP site detection is improved.

The invention relates to the molecular biology field, and discloses a specific oligonucleotide probe for detection of SNP loci of rs4244285681G / A and rs4986893636G / A of a CYP2C19 gene and a pair of specific primers for amplification of the CYP2C19 gene during detection of the above SNP loci. The probe can hybridize with different genotypes of the CYP2C19 gene specifically, and the genotypes comprise a CYP2C19*1 type, a CYP2C19*2 type and CYP2C19*3 type. The products can be used for detection of metabolic abilities for relevant medicines (such as plavix, omeprazole, voriconazole and the like) and of Chinese population, clinical medication schemes can be guided and adjusted, clinical personalized medication can be provided with a basis, the curative effects can be raised and the risks of toxic and side effects can be reduced.

The present invention relates, generally to a method of determining and assessing cytochrome P450 2C19-related (CYP2Cl9) metabolic capacity in an individual mammalian subject via a breath assay, by determining the relative amount of 13CO2 exhaled by the subject upon intravenous or oral administration of a 13C-labeled CYP2C19 substrate compound. The present invention is useful as a non-invasive, in vivo assay for evaluating CYP2C19 enzyme activity in a subject using the metabolite 13CO2 in expired breath, to phenotype individual subjects and to determine the selection, optimal dosage and timing of drug administration.

The invention belongs to the field of pharmaceutical chemistry technology, and particularly discloses thienopyridine derivatives, and preparation methods and a medical use thereof. Through structural modification of clopidogrel and prasugrel, a series of new thienopyridine derivative compounds are synthesized and mainly include derivatives esterified with ligustrazine formic acid and shikimic acid; the compounds go into a body, then are rapidly metabolized into effective metabolites and ligustrazine formic acid or shikimic acid, successfully keep away from metabolism of CYP2C19 enzyme, can be directly metabolized into active compounds to play a pharmacological function, thereby solving the clopidogrel resistance problem, effectively improving the compound antithrombotic activity, and also having no significant effect on hemorrhage risk; and the compounds have relatively ideal protective function on liver and kidney, and also have potential therapeutic significance for other cardiovascular diseases.

The invention discloses a composition for detecting CYP2C19 gene polymorphism and its application. The single nucleotide polymorphism site in the CYP2C19 gene is detected through FAM, HEX, ROX (modified) three-channel multiple PCR reaction. In order to improve the detection simplicity and specificity, one-tube parting detection is realized through the technical manner of a probe parting, thus the whole operation and the reaction process are simplified; on this basis, locked nucleic acid modification and second-grade structure modification are performed on the probe to increase the Tm value of the probe, and further improve the probe combining specificity; finally, the lowest detection limit, accuracy and the specificity of the whole kit are improved through adjusting the mixture ratio of the primer pair, dosage of Taq enzyme, dosage of magnesium ion and others; the lowest detection concentration of the kit can reach 1 mg / mu l; the accuracy and the specificity can reach 100%.

The invention relates to the field of molecular biology, discloses a method for quickly detecting CYP2C19 genetic polymorphism and a kit, and provides the method for quickly detecting the CYP2C19 genetic polymorphism and the kit. A sample to be tested is mixed with sample treating fluid, a DNA extraction step is not needed, and simple and easy treatment on samples can be realized in short time; moreover, further through a selective amplification primer and a selective detection probe, effective distinguishing between wild type variants and mutant type variants of the CYP2C19 gene can be realized, and a whole process from treatment of the samples to acquisition of a final detection result is completed within 1 hour; moreover, the accuracy, the specificity and the sensitivity of the detection can be ensured.

Primer sets for amplifying target regions containing sites to be detected in the CYP2C19 gene by a gene amplification method are provided, wherein the primer sets can amplify the regions specifically. Two pairs of primer sets are used including forward primers consisting of the base sequences of SEQ ID NOs: 12 and 32 as well as reverse primers consisting of the base sequences of SEQ ID NOs: 22 and 48, respectively. The use of these primer sets makes it possible to amplify two target regions including parts where two types of polymorphisms (CYP2C19*2 and CYP2C19*3) of the CYP2C19 gene are generated, respectively, in the same reaction solution at the same time.

Disclosed is a primer set for amplifying a target region in CYP2C19 gene which contains a site to be detected by a gene amplification method, and which enables to amplify the region specifically. Two pairs of primer sets are used, each of which comprises a forward primer comprising a nucleotide sequence depicted in each of SEQ ID NOs:12 and 32 and a reverse primer comprising a nucleotide sequence depicted in each of SEQ ID NOs:22 and 48, respectively. These primer sets enable to amplify two regions each containing a site having each of two types of polymorphisms occurring in CYP2C19 gene (CYP2C19 gene*2, CYP2C19*3) simultaneously in a single reaction solution.

The invention provides a cerebral apoplexy pharmacy and genetic correlation detection method. According to the detection method, primers and probes are synthesized through four polymorphic sites of two genes of cytochrome P450CYP2C19*2: rs4244285, CYP2C19*3: rs4986893, CYP2C19*17: rs12248560, and SVIL: rs17834991, real-time fluorescence quantification allele probe detection is implemented, and whether a target belongs to pharmacy susceptible populations or not can be judged. By adopting the method, clinical pharmacy schemes can be instructed and adjusted, bases can be made for clinical individual treatment, and adverse reactions of medicines are prevented. The method is capable of simultaneously detecting four polymorphic sites and has the advantages of being simple and convenient, accurate, rapid, high in flux and the like.

The invention discloses a quick detection kit for the CYP2C19*2 genetype. The quick detection kit comprises a PCR mixed solution, wherein the PCR mixed solution comprises the following raw materials: DNA (deoxyribonucleic acid) polymerases, DNTPs, CYP2C19*2 forward primers, CYP2C19*2 reverse primers, CYP2C19*2 mutant molecular beacons, CYP2C19*2 wild-type molecular beacons, MgC12, 5x Colorless Reaction Buffer and a cell lysis solution. The quick detection kit overcomes the defects that the conventional kit aiming at clinically detecting the CYP2C19*2 genetype and the detection method thereof cannot satisfy quick diagnosis of instructing clinical medication, and the quick detection kit and the detection method thereof provided by the invention can quickly detect the CYP2C19*2 genetype, and reduces patients' medical treatment risk.

The invention provides methods, PCR primers and sequence determination oligonucleotides for determining a human's capacity to metabolise a substrate of the CYP2C19 enzyme using genetic analysis.

The invention provides 2, 3, 5, 7-tetrasubstituted dihydro-pyrazolo piperidine derivative and a preparation method and application thereof. The derivative is 2, 3-bis(substituted phenyl)-5-subsituted arylmethyl-7-substituted benzylidene dihydro-pyrazolo piperidine derivative, having the following formula (I). The preparation method includes using substituted arylmethyl amine and methyl acrylate as raw materials; subjecting the materials to Michael addition, Dieckmann condensation and hydrolysis-decarboxylation sequentially; allowing for Aldol reaction with substituted benzaldehyde to obtain intermediate N-substituted arylmethyl-3, 5-bis(substituted benzylidene)-4-piperidone; allowing for condensation with substituted phenylhydrazine to obtain a compound according to the formula (I). The derivative is efficient in inhibiting multiplication of various carcinoma cell lines such as leukemia, esophagus cancer, ovarian cancer and breast cancer in human, is well stably metabolic in liver microsomes of human and rat, is free of direct and competitive inhibition on five enzymes of liver microsomes, such as CYP3A4, CYP2D6, CYP2C9, CYP1A2 and CYP2C19, is highly bioavailable, is low in toxicity to normal cells, and is available for the preparation of drugs for the cancers.