62 results about "Sequence specific primer" patented technology

The present invention provides a solid support which is preferably a bead comprising at least two sequence specific amplification primers wherein at least one primer is bound to the support with an inducible cleavable linker. The present invention also provides various method for preparing a solid support comprising at least two sequence specific primers, further characterized in that at least one of the primers is cleavable.

The invention belongs to the field of pharmacogenomics and genetic diagnosis, and relates to a method used for detecting HLA-B*5801 alleles. The method comprises following steps: a DNA sample to be detected is taken, three pairs of specific primers and a pair of internal primers are taken, amplification of DNA segments is realized by using sequence specific primer method, and then the results of the amplification are analyzed by agarose gel electrophoresis; or sample DNA is extracted, a pair of specific primers, a pair of internal primers and three fluorescence probes are taken, amplification of DNA segments is realized by Taqman probe method using a fluorescence ration PCR instrument, and then the amplification curve is analyzed so as to obtain results. Results analysis methods such as agarose gel electrophoresis, high resolution melting curve and SYBRGreen fluorogenic quantitative PCR are employed in the method. The method has advantages of speediness, convenience, flexibility, high resolution and no contamination; is suitable for detection of HLA-B*5801 alleles in samples such as peripheral blood and hair; and can be used for determining the probability of severe skin adverse reaction of patients with gout or hyperuricemia caused by taking of allopurinol.

The invention discloses a multiplex polymerase chain reaction (PCR) amplification method and a reagent kit for multiplex PCR amplification. The multiplex PCR amplification method comprises the step of designing public primers and multiplex primers with the public primers. The public primer design needs to follow two rules that: (1) under any optimized amplification conditions, no specific products are produced during the amplification of the public primers and templates to be measured; and (2) when the specific products are obtained through the amplification of the multiplex primers with the public primers, the public primers have the capability of amplifying the specific products, and then, a multiplex PCR system is configured for amplification. Because of the introduction of the public primers, the sequence specificity primer pair concentration in the reaction system is reduced by the level of more than four pairs, the possibility of the multiplex primers in the low concentration togenerate the mutual action in the same reaction system is greatly reduced, the problem of the technical bottle neck in the multiplex PCR of mutual interference of different primer pairs can be solved, and the precondition can also be created for adding new primer pairs and increasing the detection gene number.

The present invention relates to the fields of biotechnology and medicine, and provides specific sequence specific primers-polymerase chain reaction primer sequences and a kit for simultaneously detecting human MTHFR gene polymorphism and human MTRR gene polymorphism, wherein the kit contains the specific primers, specific probes, an internal standard system, Taq enzyme and UNG enzyme. According to the present invention, the primers and the kit have advantages of strong specificity, high sensitivity, simple and rapid operation, high-throughput, safety, objective result interpretation, and the like.

The invention discloses a detection kit and a detection method for detecting SNP sites related to individualized drug use of tacrolimus and cyclosporine A by using multiplex PCR technology combined with SNP sensitive molecular switch technology. The kit is used to type three SNP loci associated with the administration of tacrolimus and cyclosporine A, including: rs2242480 SNP locus on the CYP3A4 gene, rs776746 SNP locus on the CYP3A5 gene, and rs776746 SNP locus on the MDR1 gene rs1045642 SNP site. The kit contains wild-type 2× amplification buffer, mutant 2× amplification buffer, and polymerase. The two buffers contain sequence-specific primers and internal reference primers corresponding to the SNP wild-type and mutant phenotypes, which can The typing of the above three SNP sites was completed in two multiplex PCR reactions, so as to provide a molecular biological basis for the rational use of tacrolimus and cyclosporine A.

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 discloses a detection kit for detecting SNP sites related to homocysteine metabolism by using multiplex PCR technology and SNP sensitive molecule switch technology and an amplification method and a detection method thereof. The kit carries out genetyping on four SNP sites related to homocysteine metabolism, i.e., an rs1801133 SNP site in the MTHFR gene, an rs1805087 SNP site in the MTR gene, an rs1801394 SNP site in the MTRR gene and an rs5742905 SNP site in the CBS gene. The kit comprises a wild 2* amplification buffer, a mutant 2* amplification buffer, polymerase, a cell lysis solution and glycerin, wherein the two buffers respectively contain sequence-specific primers and internal control primers corresponding to SNP phenotypes consisting of a wild type and a mutant type, genetyping of the above-mentioned SNP sites can be finished in two multiplex PCR reactions, so homocysteine metabolic capability and demands for folic acid, VB12 and VB6 of a subject are known.

The invention belongs to the technical field of genetic engineering, relates to a primer combination, a kit and a method for performing PCR-SSP (polymerase chain reaction-sequence specific primer) typing detection on a KIR (killer cell immunoglobulin-like receptor) gene. The primer combination consists of detection primers shown in SEQ ID NO. 1-65, wherein detection primers 2DL1 and 2DL2 form a group, 2DS3 and 2DP1 form a group, DRB1, 3DS1 and 3DL1 form a group, 3DP1, 2DL5 and 2DS1 form a group, 2DL3 and 2DL4 form a group, 2DL3, 2DS3 and 3DS1 form a group, 2DS4, 3DL1 and 2DL1 form a group, 2DS4 and 2DS5 form a group, 3DL3, 2DS5 and 3DL2 form a group, 2DP1 and 2DS2 form a group, 2DS2 forms a group, 2DL4, 3DL2 and 3DL3 form a group, and all the detection primers form 12 primer groups. Through the primer combination, the kit and the method, the accuracy and the reliability are high, the operation is simple and stable, and a result is easily judged.

The invention relates to a primer group, a method and a kit for detecting the triploids of a fetus by virtue of plasma free DNS in maternal peripheral blood, wherein the DNA is selected from human No. 21, No. 18 and No. 13 chromosomes, 300 pairs of sequence specific primers are designed with regard to the No. 21, No. 18 and No. 13 chromosomes respectively, each pair of the sequence specific primers comprises a forward primer, a reverse primer and a middle primer, and the gene sequence numbers of the primers are SEQ ID NO: 1-2700. The primer group, the method and the kit disclosed by the invention have the following beneficial effects: the triploids of the fetus are detected by the method disclosed by the invention, the detection is non-invasive detection without a threat on the health of the fetus, diagnosis and detection are high in accuracy and low in expense, flow and data analysis are simple and convenient, and PCR reaction condition requirements are not rigorous; the primer group, the method and the kit are wide in applicability, can be used for promoting the popularization of non-invasive screening for a trisomic syndrome, and are conductive to improving the population quality of China.

The invention discloses a method and a kit for detecting drug resistance of mycobacterium tuberculosis. The invention provides special-purpose primers and special-purpose probes for identification of drug resistance of mycobacterium tuberculosis in a sample needing to be detected, wherein the special-purpose primers comprise multiple primer groups; each one of the primer groups comprises a commonprimer and multiple specific primers for amplification of alleles corresponding to all mutational sites of same-purpose genes; the special-purpose probes comprise multiple types of probes; and each one of the probes specifically binds with target sequences corresponding to the common primer and the specific primers of the primer group. Experiments of the invention prove that the special-purpose primers and the special-purpose probes can be combined by an efficient and sensitive duplex Taqman real-time fluorescent polymerase chain reaction (PCR) technology and a sequence specific primer (SSP) amplification technology so that drug resistance of mycobacterium tuberculosis in a detected sample can be detected in a short time.

We provide a DNA sequencing method and a sequencing system where large numbers of sequence reads can be obtained in parallel by running traditional electrophoresis in a special format. Parallelization is obtained either through a 3-dimensional gel-cube or through bundled capillary tubes including fiber-optic tubes or other types of micro channels in a bundle or matrix format. Various ways of capturing sequence traces are provided. We also provide two distinct methods for preparing genomic DNA / cDNA fragments: one through universal primer site anchoring and amplification of single molecules, and the other through micro-array / bead oligomer extension and dye-terminator incorporation using target sequence specific primers. The invention can perform large-scale genomic sequencing including sequencing a complete human genome in one or a few runs.

The invention belongs to the field of biotechnology, particularly relates to a titer detection method for a slow virus carrier, and discloses a quality index detecting method conducting real-time fluorescence quantification nucleic acid amplification detection (qPCR and RT-qPCR) by using a sequence specific primer and a fluorescent dye, a used primer and a standard product. The detecting method isefficient and accurate, the operation is easy, the use amount of a sample is low, the repeatability is good, and the limit problem of inaccurate carrier quantitation in the fields of eucaryon geneticengineering and gene therapy is solved. The method has the advantages of detection time and simple and convenient operation step, the analysis process after the amplification is not needed, comparedwith the Taqman probe method, the SYBR Green I does not need the design of a synthesis fluorescent probe, the design is simplified, the cost is lowered, and a dissolution curve is combined in the analysis to judge the specificity of the reaction.

The invention relates to a reagent used for detecting antigen genotypes relevant to the anaphylactic reaction of antiepileptic drugs, and a clinical application method thereof. The reagent is characterized in that: 1) the reagent comprises Taq enzyme, a chain of seven tubes and a PCR primer; 2) the PCR primer is shown in the sequence list SEQ ID No.1. The clinical application method is characterized in that: 1) four pairs of the sequence specific primers and two pairs of internal reference primers are designed by utilizing a sequence specific primer PCR-SSP according to an HLA-B sequence, and PCR amplification is carried out by taking gDNA extracted from human peripheral blood or other issues as a template; 2) gel electrophoresis is adopted for the detection so as to confirm that the genotype of the sample is HLA-B multiplied by 1502. The reagent and the method of the invention have the advantages of simple operation, high accuracy and low cost, and are especially suitable for determining that whether the antiepileptic drugs such as carbamazepine, etc. can be taken by the HLA-B multiplied by 1502 genotype detection before patients in China or Asia take antiepileptic drugs such as the carbamazepine, etc.

We provide a DNA sequencing method and a sequencing system where large numbers of sequence reads can be obtained in parallel by running traditional electrophoresis in a special format. Parallelization is obtained either through a 3-dimensional gel-cube or through bundled capillary tubes including fiber-optic tubes or other types of micro channels in a bundle or matrix format. Various ways of capturing sequence traces are provided. We also provide two distinct methods for preparing genomic DNA / cDNA fragments: one through universal primer site anchoring and amplification of single molecules, and the other through micro-array / bead oligomer extension and dye-terminator incorporation using target sequence specific primers. The invention can perform large-scale genomic sequencing including sequencing a complete human genome in one or a few runs.

The invention relates to a multi-PCR method capable of detecting a human RhD blood type and gene type and an in-vitro diagnosis kit prepared using same, which can detect the human RhD blood type and gene type comprising gene types of D / D, D / d, DEL, DEL / d and d / d. The method is finished by utilizing the change of the specific sequence sites of different gene types of the human RhD blood type, designing a plurality of pairs of primers and carrying out a polymerase chain reaction (PCR). Thereby, the invention can be regarded as being established on the basis of the principle of PCR-SSP (sequence specific primer). By utilizing the sequence specific primer designed aiming at different sites and optimizing the mixing proportion, the reaction buffer components and the PCR reaction conditions of the sequence specific primer, the purpose of accurately detecting the human RhD blood type and gene type is achieved. In one implementation, the invention provides the multi-PCR method for detecting the human RhD blood type and gene type; and in another implementation, the invention provides the kit for detecting the human RhD blood type and gene type. The details of one or a plurality of implementations of the invention are described in the accompanying drawing and the specification. By reading the appended drawings, the detailed descriptions and the claims, a user can clearly learn about other characteristics, purposes and advantages of the invention.

The invention relates to a multiplex polymerase chain reaction (PCR) method capable of detecting human RhD blood type genotypes and an in vitro diagnostic kit prepared by applying the method and capable of detecting the human RhD blood type genotypes comprising D / D, D / d, DEL, DEL / d and d / d genotype. According to the method, variation of specific sequence loci of the different genotypes of a human RhD blood type is utilized to design a plurality of pairs of primers to perform a PCR. Accordingly, the multiplex PCR method and the in vitro diagnostic kit can be regarded to be on the basis of a PCR-SSP (sequence specific primer) principle. By designing the sequence specific primers according to different loci and by optimizing mixing proportion, reaction buffer solution ingredients and PCR reaction conditions of the sequence specific primers, the aim of accurately detecting the human RhD blood type genotypes is achieved. In an embodiment, the multiplex PCR method for detecting the human RhD blood type genotypes is provided, and in another embodiment, the kit for detecting the human RhD blood type genotypes is provided. Details of one or a plurality of embodiments are described in attached drawings and explanations. By reading the attached drawings, detailed descriptions and claims, other characteristics, aims and advantages of the multiplex PCR method and the kit can be learned about clearly.

A cost-effective multi-channel capillary gel-electrophoresis system for highly efficient, nucleic acid-based analysis, in particular HLA SSP (Human Leukocyte Antigen Sequence Specific Primer) typing applications. Twelve DNA samples are automatically injected, electrophoretically separated, detected and analyzed simultaneously by using a multiple usage and disposable multi-capillary gel-cartridge. Using commercially available DNA size markers as indicators, the system provides high resolving power with 12-channel separations in less than 1.5 minutes. The system can hold a total of 96 samples in a PCR plate that can automatically be analyzed within 25 minutes for a full plate of the HLA SSP kits.

The present invention provides a solid support which is preferably a bead comprising at least two sequence specific amplification primers wherein at least one primer is bound to the support with an inducible cleavable linker. The present invention also provides various method for preparing a solid support comprising at least two sequence specific primers, further characterized in that at least one of the primers is cleavable.

The invention discloses a method of detecting common ambiguous types of HLA (human leukocyte antigen) typing locus A. The method includes: (1) acquiring ambiguous types of locus A through HLA typing results, and selecting specific primers necessary to distinguish the ambiguous types, wherein the specific primers are shown as SEQ ID NO. 1 to 9; (2) using the selected specific primers to perform sequencing PCR (polymerase chain reaction) on the purified HLA-A gene amplification product; (3) purifying the post-sequencing product, acquiring a product sequence through a sequencer, comparing the product sequence with the ambiguous types so as to distinguish the ambiguous types, and finally determining HLA-A type information. Compared with traditional PCR-SSP (PCR-sequence specific primer) methods to determine gene ambiguous types, the method herein can accurately and precisely judge the types, has greatly reduced cost, is simple to perform, can easily provide high throughput, and is a time-saving economical option for large-scale donor HLA typing.

The invention discloses a detection reagent and a typing method for the simultaneous genotyping of antigens of ABO blood group and specific A2, comprising four primers, two of which are sequential specific primers and the other two of which are inner comparison primers. The method can carry out simultaneous amplification to the blood group genes of the antigens of the ABO blood group and the specific A2 subtypes, which are very common among the Chinese population. The method can detect four alleles, A101, A102, B101 and O01, and six A2 alleles, A201, A202, A202, A204, A205 and A206, which are very common among the Chinese population. Compared with the traditional detection methods, the detection method of the invention is fast, simple, practical, specific and sensitive, is suitable for the genetic identification of ABO blood groups of Chinese Han population and can be used in the research of the ABO blood group of human beings.

The invention relates to a rapid bidirectional multilocus gene mutation method. According to the method, by taking an improved overlap extension PCR method as a basis and adopting sequence specific primers with mutation sites, DNA bidirectional multilocus gene mutation is realized by carrying out two PCR amplifications and joining DNA segments. The method can be widely used in target gene site-directed mutagenesis, gene synthesis, protein structure-function research, protein expression optimization and target gene packaging. According to the method, three DNA segments and three corresponding pairs of specific amplification primers are designed based on sequence characteristics of DNA segments needing mutation. Target gene segments for introducing needed mutation sites are formed by carrying out two PCR amplifications and setting specific amplification programs.

The invention discloses a reagent and detection method capable of specifically detecting Mur antigen gene, the reagent is composed of 2 *premixed liquids, a Mur upstream primer, a Mur downstream primer, an internal reference upstream primer, an internal reference downstream primer, and DNA enzyme / RNA enzyme-free water, when in use, blood genome and the reagent are mixed for PCR (polymerase chain reaction) amplification reaction, PCR products are identified by electrophoresis for obtaining detecting results. According to the reagent and detection method capable of specifically detecting the Mur antigen gene, a simple, rapid, sensitive and specific PCR-SSP (polymerase chain reaction-sequence specific primer) method for detection of the Mur antigen gene in Miltenberger blood type is established by use of the Mur antigen gene specific primer, the method is accurate in detecting results, is conducive to large-scale and automated screening of the Mur antigen, can provide effective reference for clinical Mur antigen detection and identification and blood matching work, and ensures the safety of blood transfusion.

The invention provides a method for detecting medicine-resistance mutation of tyrosine-methionineaspartate-aspartate (YMDD) of hepatitis B virus, which detects the medicine-resistance mutation of tyrosine-methionineaspartate-aspartate (YMDD) of hepatitis B virus by using a real time-amplification refractory mutation system-quantitative polymerase chain reaction (RT-ARMS-q PCR) process. In the invention, sequence specific primers and a fluorescent dye SYBRGreenI are used for amplification, the real-time monitoring of a reaction is realized, the detection time is shortened, operation steps are simplified, an analysis process after amplification is not needed, and the possibility of product pollution is lowered; and compared with a Taqman probe requiring a double marking process, the SYBRGreenI does not need designing and synthesizing a fluorescent probe, the design concept is simplified, and the cost is low. And by combining fusion curve analysis, reaction specificity is realized.

The invention relates to a specific sub sequence for separating an Ac / Ds transposon flanking sequence in a corn Ac / Ds transposition mutant and a separation method thereof. The sub sequence is base sequences shown in SEQ ID NO:1 to SEQ ID NO:4. The Ac / Ds flanking sequence in a genome can be well amplified and obtained by using an Ac specific primer and a specific primer of the sub sequence after these sub sequences are connected with corresponding Ac / Ds transposon deoxyribonucleic acid (DNA) subjected to restrictive incision enzyme digestion. The detection method is simple to operate, and important technical means is provided for large-scale obtaining of the Ac flanking sequence of the Ac mutant and separation of the gene of an Ac insertion mutant.

Reagents and methods for the fast, accurate and early detection of infections in thalassemia major patients. Reagents include a primer pool containing a mixture of primer pairs for the specific recognition and simultaneous amplification of targeted gene sequences of pathogens from a biological sample collected from a patient in a multiplex PCR reaction. Pathogens may include at least one Fungi species, Klebsiella pneumoniae, Candida albicans and Staphylococcus aureus. Subsequent identification of the pathogens is achieved by DNA sequencing analysis.

The present invention provides a method for genotyping alleles in at least one homologous genetic loci set, comprising: (i) providing a DNA-containing sample that includes said at least one homologous genetic loci set; (ii) performing PCR amplification of regions of said homologous genetic loci set using consensus sequence-specific primers, wherein said consensus sequence-specific primers bind to consensus sequences that are common to a plurality of genes within the genetic loci set, thereby generating a pool of amplification products; (iii) sequencing a plurality of said amplification products in order to determine the relative proportion of each nucleotide at each position in a sequencing read; (iv) performing a sequence alignment between the sequencing read results of (iii) and at least one reference sequence, which reference sequence corresponds to one of the genes in said homologous genetic loci set; and (v) performing genotype calling of the allele or alleles in said sample based on the relative proportion of each nucleotide at each of a plurality of discriminant positions in said alignment. Also disclose are related products, kits and systems for performing the method.

The invention belongs to the technical field of molecular biology, and discloses an ITS sequence, a specific primer pair, an identification kit, and an identification method used for identification ofdendrobium officinale and high imitation fake products. The ITS sequence possesses a nucleotide sequence represented by SEQ ID NO:1, and can be used for identification of dendrobium officinale and high imitation fake products. The kit used for identification of dendrobium officinale and high imitation fake products is also provided; establishment of a related PCR detection system is carried out,molecular identification basis is provided for dendrobium officinale medicine material identification, rapid accurate detection and distinguish of dendrobium officinale from dendrobium officinale medicine materials with relatively high similarity are realized. Operation is simple and convenient; the specificity is high; the sensitivity is high; detection on dendrobium officinale medicine materialsis realized; and the identification method is suitable for wide popularization and applications.

The invention discloses a method for detecting human HLA-B*5801 alleles. The method comprises the following steps: 1) designing a sequence-specific primer to search a DNA sequence of a target gene; screening out proper primer pairs in accordance with detection sites of the target gene; and combining the various sites, so that various PCR reaction systems are defined; 2) extracting genome DNA of human peripheral blood cell samples, and conducting amplification via primers which are screened out; 3) conducting SAP treatment on amplification PCR products, implementing single base extension and conducting purifying, so that sample analytes are obtained; and 4) applying the sample analytes to a chip matrix, and implementing analysis via matrix-assisted laser desorption ionization time-of-flight, so that sample molecular weights are obtained; and detecting out corresponding site information of the alleles, thus completing genetic typing analysis. The method provided by the invention has theadvantages that 384 samples can undergo multiple detection via one chip, each system can accept 30 multiples of reactions to the greatest extent, and multiple HLA-B*5801 allele sites can be simultaneously detected.

The invention provides a primer set for detecting genotyping of acetaldehyde dehydrogenase 2, a reagent, a kit, a detection method and application, and relates to the technical field of genetic engineering. The primer set for detecting genotyping of acetaldehyde dehydrogenase 2 comprises a common upstream primer, an allele-specific primer, and an intermediate sequence-specific primer. The primer set has the characteristics of low development cost, short development cycle, high accuracy and the like, and can meet the requirements of accuracy, precision and detection rate and the like required for clinical detection. Moreover, the primer set has the advantages of high specificity, high accuracy, and wide range of DNA measurement, and is favorable for promoting the clinical detection of the nitroglycerin individualized drug gene for the locus. The method for detecting genotyping of acetaldehyde dehydrogenase 2 provided by the invention has low detection cost, and the genotype of ALDH2 rs671 locus can be analyzed economically, accurately and rapidly by using the detection method provided by the invention, which is of great significance in clinical promotion of the nitroglycerin individualized drug gene detection.

The invention discloses a PCR (Polymerase Chain Reaction) primer group for detecting pepper mild mottle virus, the PCR primer group comprises three pepper mild mottle virus (PMMoV) genome motor protein (MP) and coat protein (CP) coding gene partial sequence specific primers and two pepper reference gene Actin specific primers, wherein the three pepper mild mottle virus (PMMoV) genome motor protein(MP) and coat protein (CP) coding gene partial sequence specific primers are PMMoV-F1, PMMoV-F470 and PMMoV-R991 respectively, and the partial sequence specific primers of the MP gene and the CP geneare PMMoV-F470 and PMMoV-R991; the two pepper reference gene Actin specific primers are PeActin-F1 and PeActin-R1093. The PCR amplification result is detected in 1.0%-1.5% agarose, an expensive fluorescent quantitative PCR instrument is not needed, and the PCR primer group has the advantages of being rapid, economical, high in stability, high in accuracy and high in sensitivity.