1853 results about "Fluorescent pcr" patented technology

The invention relates to a microfluidic control chip which is formed by bonding and sealing PDMS (Polydimethylsiloxane) material integrated with a micro valve, and comprises an upper layer of air control passage, a middle layer of PDMS membrane and a lower layer of microfluidic passage. A real-time PCR virus quick detection device comprises the microfluidic control chip, a temperature control unit, a signal detection unit and a data processing and control unit. The invention designs the microfluidic control chip which is integrated with the micro valve, is in a three-layer PDMS structure, andis used for virus detection and the real-time fluorescent PCR detection device. Compared with the traditional method, the invention has the advantages of short time, less sample usage, quick detection speed, simpleness and convenience in operation, integration and the like.

The invention discloses a rapid fluorescence PCR detection kit for ASFV (African swine fever virus). The kit comprises specific primers ASFV-F and ASFV-R as well as a TaqMan probe ASFV-P, the 5' end of the probe labels fluorescence dye as FAM and the 3' end labels a fluorescence quenching group as BHQ-1. The kit can be used for detecting the ASFV in nasal swabs, blood, serum, plasma and tissue ofswine to realize rapid detection of the ASFV. In the whole ASFV detection process, only 40 min is taken from DNA extraction to obtaining of detection results, so that the detection time is greatly shortened, and the detection efficiency is improved.

The invention relates to a biological detection reagent capable of discriminating three types of ruminant animal varieties simultaneously, a preparation method and an application thereof. The invention selects the specificity of the cow, goat and sheep, and the conservative sequence segment of conservative mitochondrial gene as target, applies Primer Eexpress 3.0 software and Primer Select software in DNAStar, and designs and synthesizes a plurality of groups of primers and probes. After being synthesized and marked, the plurality of pairs of designed primers and the probes are carried out best pairing screening experiment so as to obtain the fittest combination of primers and probes. The reagent contains two pairs of specificity primers, and three Taq Man probes, wherein one pair of theprimer aims at cow source components, the other pair of primer aims at common primers of goats and sheep source component detection; the three probes are respective source components to the cows, thegoats and the sheep; and the amplified target fragment length to the source component detection of the cows, the goats and the sheep is respectively 92bp, 136bp and 136bp. The triple Taq Man fluorescence PCR detection method can simultaneously carry out accurate quantitative detection to the source components of the cows, the goats and the sheep, has simple and convenient operation, intuitive result, strong specificity, high sensitivity and good repeatability and can realize high throughput detection of various source components.

The invention relates to a method utilizing a fluorescence PCR technology to identify mouse genotype. A fluorescence PCR primer is utilized, after PCR amplification, the amplification product is mixed with ROX with a known size, the size of PCR products can be calculated by Genemapper software through 377 sequencing; and thus the mouse genotype can be identified. The provided method can simply, directly, and precisely find out the mice having micro RNA, whose Crispr/Cas9 has been knocked off.

The invention discloses a primer and probe for detecting a human epidermal growth factor receptor (EGFR) gene and/or a K-ras gene, a kit containing the primer and the probe and a device for detecting genetic mutation on the basis of a digital PCR platform.The method for detecting the genetic mutation by means of the primer and the probe comprises the steps that the prime and the probe are provided; DNA of a sample to be detected is extracted; a fluorescent PCR reaction system capable of amplifying a mutant gene sequence is prepared; a target probe and an internal reference probe are utilized to be hybridized with amplified products respectively, and fluorescent signals of corresponding fluorescent groups are detected; existence of the genetic mutation is judged and/or the mutation rate is calculated according to the strength and proportion of the fluorescent signals of the target probe and the internal reference probe.According to the method for detecting the genetic mutation, the needed primers and probes are small in number, the optimization procedure is simple, related mutation of EGFR and/or K-ras gene can be qualitatively or quantitatively detected, and the detection sensitivity is high; a DNA sample with low initial amount can also be detected stably.

The invention relates to a primer middle sequence interference PCR (Polymerase Chain Reaction) technology. The improved PCR technology is characterized in that one segment of relatively non-complemented or same-sequence basic group primer molecules in the intermediate domain of primers perform the antisense interference inside and outside so as to competitively destroy the polymerization among the primers to selectively inhibit the primer dimer (PD) from being amplified. For the interference of the intermediate domain of the primers, based on the primers optimally selected by the conventional design principle, the technology that the intermediate domain (ID) of a pair of the primers are in parallel but are not complemented with each other or are in the same sequence or/and the technology that ID antisense oligonucleotides (Oligo) are added into the primers to perform the interference action or/and the Oligo antonymy is carried out in the primer molecules via the ID so as to perform the interference action are adopted, or the combined technology of the three types of the technologies is adopted. As a result, only the ID of the primers is interfered while the target specific amplification is not influenced; the combining force acting on the minority of base-group pairing hydrogen bonds at the tail end and the base-group hydrogen bonds outside the tail end due to the action of the primers is dispersed to a maximum extent, so that the PD is selectively inhibited. Therefore, the PD accumulation in the PCR system is avoided. If the mineral oil is additionally used, the sealed primers can slowly release the hot starting and the UDG pretreatment so as to prevent aerosol glue as a byproduct of the PCR system from causing the pollution. Consequently, the nucleic acid is amplified reliably and the real-time fluorescence PCR is quantified accurately.

The invention provides a method for single-tube multiplex fluorescent polymerase chain reaction (PCR) detection of human coronavirus OC43, 229E, NL63, HKU1 and SARS. The method adopts primers having sequences of SEQ ID NO: 1-10, and probes having sequences of SEQ ID NO: 11-15. The invention also provides a kit for single-tube multiplex fluorescent PCR detection of human coronavirus OC43, 229E, NL63, HKU1 and SARS. The kit contains the primers and the probes. The method provided by the invention adopts the primers which are specific primers of OC43, 229E, NL63, HKU1 and SARS, and the probes which are Taqman probes, utilizes TAMRA/CY5/FAM/ROX/JOX multiple fluorescein labeling, realizes single-tube multiplex detection of human coronavirus OC43, 229E, NL63, HKU1 and SARS, and has the advantages of strong specificity, high sensitivity, fast detection speed, simple and convenient operation and low cost. The primers and the probes can be used as detection reagents for a scientific research and clinical application.

The invention relates to a real-time fluorescence PCR (polymerase chain reaction) detection system based on a rotary type microfluidic chip. The system comprises an annealing low-temperature area, an extension appropriate temperature area, a thermal change high temperature area, a rotating module, a PCR reagent accommodating cavity, an injection micro-channel, an instrument fixing base and a fluorescence detection system, wherein a temperature cycle control module comprises the annealing low-temperature area, the extension appropriate temperature area and the thermal change high temperature area; a micro-channel cycle PCR amplification module comprises the rotating module, the PCR reagent accommodating cavity, the injection micro-channel and the instrument fixing base; and a fluorescence spectrum detection module comprises the fluorescence detection system. According to the system, three modules, namely, the micro-channel cycle PCR amplification module, the temperature cycle control module and the fluorescence spectrum detection module are integrated, so that the portable micro PCR fluorescence real-time detection system applicable to space operation requirements is realized, and purposes of function integration, structure microform, lightness, small size and full-automatic detection are achieved.

The invention provides a kit for jointly detecting respiratory tract pathogen through a multiple fluorescent PCR method. The kit comprises six components: reaction liquid A, reaction liquid B, reaction liquid C, enzyme mixed liquid, positive control and negative control, and comprises 11 common respiratory tract pathogen detections (general type of influenza virus A, influenza virus B, respiratory syncytial virus, 1/2/3 type of human parainfluenza virus, adenovirus, mycoplasma pneumoniae, chlamydia pneumonia, legionella pneumophila, streptococcus pneumonia, haemophilus influenza, A streptococcal); the amplification is performed through three reaction buffers, and each reaction buffer contains four fluorescent channels, 90% pathogen infection on the clinic can be checked.

The invention relates to analysis of the MSI (microsatellite instability) state which can serve as an auxiliary diagnosis marker for tumor immunotherapy (PD-1/PD-L1). The MSI of tumor cells of human beings is detected on the basis of a fluorescent PCR platform, and five of more mononucleotide repeat loci (NR-21, NR-24, BAT-25, BAT-26 and MONO27) reported in documents and internationally are selected for analysis of the MSI state. Besides, two pentanucleotide repeat loci (Penta C and Penta D) are selected for quality control of samples and are mainly used for detecting mixing and pollution of the samples. A fluorescently labeled primer is adopted, and besides, 7 loci are amplified; by comparison with other products, the amplification conditions are simplified, amplification can be performed on common PCR equipment, signals are stronger in combination with capillary electrophoresis analysis, the operation is simple, economical and practical properties are realized, the MSI state of a patient who receives or is going to receive tumor immunotherapy (PD-1/PD-L1) can be detected within short time, and guiding significance is given as soon as possible for clinical treatment.

The invention discloses a method for detecting an allergen almond component in foods by a fluorescent PCR technology, belonging to allergen detecting technologies, in particular a method for detectingan allergen almond component in foods by an exonuclease probe fluorescent PCR technology (TaqMan). Aiming at an allergen almond component Pru du 1.06B DNA sequence a primer and a TaqMan probe are designed, the fluorescent PCR detecting method is established. The method includes the designed primer and a probe of the almond component specificity, and the fluorescent PCR reaction condition matchedwith the primer and the probe. The method has no cross reaction with peanuts, hazelnuts, chestnuts, walnuts, pine nuts, macadamia nuts, and the like and has specificity; and the detecting sensitivitycan reach 5mg/kg. The method can be used for detecting the allergen in the foods and preventing anaphylactic reaction caused by the foods and has practical meanings.

The invention relates to a micro-fluid fluorescence speed measurement and control device with a fluorescence PCR micro-fluidic control chip and a method thereof. The invention belongs to the detection field of biology, analytical chemistry and medicine. The invention comprises an illuminant (1), an exciting light splitting system (2), an emitted light splitting system (4), a flow speed regulation and control actuator, a photoelectric detector connected with a computer (3), a lead screw and a stepping motor connected with the lead screw. A check point is arranged at the same position of each Microchannel of a biologic PCR fluorescence microfluidic control chip, respectively. An exciting light optical fiber conducting system and an emitted light optical fiber conducting system are fixed on the lead screw. The computer controls the movement of the lead screw through the stepping motor; the input end of the speed flow regulation and control actuator is connected with the computer; the output end of the flow speed regulation and control actuator is connected with the Microchannel of the biologic PCR microfludic control chip. The device can measure the flow speed of each Microchannel, and the computer can adjust and control the flow speed of the next Microchannel through the flow speed regulation and control actuator according to the difference between the actual speed and the theoretic speed.

The invention provides a freeze-drying method for a fluorescent PCR (Polymerase Chain Reaction) amplification reagent and application of the fluorescent PCR amplification reagent. The fluorescent PCRamplification reagent is prepared from a fluorescent PCR amplification buffering solution, a primer, a probe, a hot start Taq enzyme, a UNG enzyme, dATP, dGTP, dCTP, dTTP, dUTP and a freeze-drying protection agent. All components of the fluorescent PCR amplification reagent are uniformly mixed; after a mixture is pre-frozen at -35 DEG C for 3h, air pressure of a freeze-drying machine is reduced tobe 10Pa or lower, and then the mixture subjected to vacuum treatment for 2h at -35 DEG C, vacuum treatment for 2h at 10 DEG C and vacuum treatment for 2h at 30 DEG C, so as to obtain fluorescent PCRamplification reagent freeze-dried powder. According to the freeze-drying method provided by the invention, all the components needed by fluorescent PCR are prepared into the same freeze-dried powder,and room-temperature transportation and preservation can be realized; the freeze-dried powder can be dissolved by only utilizing a freeze-dried powder dissolving solution in a utilization process andthen is added into a sample. The freeze-dried powder has the characteristics of convenience for utilization, stability and reliability.

The invention discloses a fluorescence quantitative PCR (polymerase chain reaction) universal premier for detecting aspergillus, detection probes and a detection kit, wherein the detection kit comprises universal premiers displayed by base compositions such as SEQ ID. NO: 1 and SEQ ID. NO: 2; and the detection probe is CY5-TAAAGTTGGGTGTCGGCTGG-BHQ aiming at aspergillus fumigatus, the detection probe is FAM-TTGATTTGCGTTCGGCAAGC-BHQ aiming at aspergillus flavus, the detection probe is HEX-ACAAGTTGCAAATAAATGCGTCG-BHQ aiming at aspergillus, and/ or the detection probe is ROX-ATGGTTGGAAAACGTCGGCA-BHQ aiming at aspergillus Niger. The multiple PCR detection method, premier, the probes and the kit thereof provided by the invention aim at four main pathogenic aspergilli and have high specificity and sensitivity; and the detection method is rapid, simple and convenient, and can be used for detecting and identifying the strain of pathogenic bacteria.

A provided multiple fluorescence PCR detection kit for clostridium difficile toxin genes mainly comprises specific primers, probes and a PCR reaction reagent, and the specific primers and the probes respectively consist of specific primers and probes of clostridium difficile toxin A (tcdA), toxin B (tcdB), binary toxin A (cdtA) and binary toxinB (cdtB). The beneficial effects of the invention comprise: the fast, sensitive and specific multiple fluorescence PCR detection kit and a detection method are provided for the clostridium difficile toxin genes, and a foundation is provided for distinguishing between toxigenic strains and avirulent strains of clostridium difficile, and early diagnosis on infection of clostridium difficile.

The invention discloses a composition for detecting the instability of a microsatellite and application of the composition. The composition comprises primer pairs respectively used for detecting specific target genes relevant with the instability of the microsatellite in a to-be-detected sample. A kit for detecting the instability of the microsatellite is used for detecting microsatellite distribution conditions of nine loci of Bat 26, Bat 25, NR-21, NR-24, NR-27, Mono-27, D5S346, D2S123 and D17S250 and three loci of Penta C, Penta D and Amel by virtue of a specific multi-fluorescence PCR fragment through an analysis capillary electrophoresis method and detecting whether the microsatellite is in a stable or instable state. By adjusting the proportion of 12 pairs of primers, the use amounts of Taq enzyme and magnesium ions, a PCR system and the like, and the sensitivity and specificity of the whole kit are improved.

The invention belongs to the technical field of molecular biology and provides primers, a probe composition and a kit for rapid identification of nine animal origin ingredients in food or feed, a detection method for identification of the nine animal origin ingredients in the food or feed and application of the primers, the probe composition, the kit and the detection method. Origin ingredients of the food or feed containing multiple species can be identified rapidly by the primers, the probe composition and the real-time fluorescent PCR (polymerase chain reaction) joint detection kit. The detection method includes: designing a universal primer by taking 16SrDNA as a target gene, and designing specific probes of nine species by designing to construct internal amplification control and designing the specific probes aiming at internal control sequences; performing PCR amplification by three PCR reaction systems; interpreting the origin ingredients of the nine species directly through corresponding fluorescent probe signals and Ct values. The detection method is low in cost, time saving and high in efficiency and can achieve identification of multiple species simultaneously.

The invention provides a micro-fluidic chip reagent kit for detecting ten respiratory tract infection pathogens and a use method of the reagent kit. The reagent kit adopts a combination of a Taqman probe fluorescent PCR (polymerase chain reaction) technology and a micro-fluidic chip, detects the ten common respiratory tract infection pathogens, can obtain a detection result within 2h, and is highin specificity, and the sensitivity can reach 100 copies/microliter. The kit comprises a sample introduction chamber, at least ten reaction chambers and a micro-fluidic flow channel, wherein the reaction chambers are mutually independent; each reaction chamber is provided with a reagent dry powder for amplifying one respiratory tract pathogen in advance; each reagent dry powder comprises a primerfor amplifying the corresponding respiratory tract pathogen and a TaqMan probe; the reagent dry powder arranged in each reaction chamber in advance can amplify any one of the ten respiratory tract pathogens; the respiratory tract pathogens amplified by the reagent dry powder in all the reaction chambers can include the ten respiratory tract pathogens.

The invention provides a Zika virus fluorescent PCR detecting kit, and relates to a kit for detecting Zika virus, in particular to simultaneous detection of Zika virus RNA by using a real-time fluorescent polymerase chain type reaction technology.The kit mainly comprises RT-PCR reaction liquid, primer probe mixed liquid, an RT-PCR reaction enzyme system, DEPC H2O and a packaging box for separating and packaging reagent bottles or tubes in a centralized manner.The kit adopts a Taqman probe detection mode and a one-step real-time fluorescent PCR reaction mode, can accurately and quickly detect the Zika virus RNA, and can be widely applied to various fields such as Zika virus disease clinic early diagnosis, disease prevention and scientific research.

The invention relates to a kit for performing gene detection on middle east respiratory syndrome coronaviruses, in particular to a kit for detecting middle east respiratory syndrome viruses UPE and N genes through multiple real-time fluorescent polymerase chain reaction technologies. The kit mainly comprises an RT-PCR solution, a primer probe mixed solution, an RT-PCR enzyme system, DEPC H2O and a packaging box for packaging reagent bottles or tubes in a separated and concentrated mode. Due to the fact that the mode that various fluorescent channels are used for detection respectively is adopted for the kit, a one-step method real-time fluorescent PCR mode is applied, the middle east respiratory syndrome coronaviruses UPE and the N genes can be accurately and fast detected, and the kit can be widely applied to various fields of cloinical early-stage diagnosis, epidemic situation prevention and control, scientific research and the like of a middle east respiratory syndrome.

The invention provides a rotary scanning real-time fluorescent quantitative PCR (Polymerase Chain Reaction) detection system. The detection system comprises a fluorescent detection device, a light source stimulation device, a thermal cycling module device and a signal transmission device. An output shaft of a stepping motor is connected with a light path gating rotary piece by an optical fiber fixing disc and a light path gating rotary piece protection ring; a base of the fluorescent detection device is provided with a hole corresponding to a detection window of a PMT protection device so that the detection window of the PMT aligns to an optical fiber output end focusing device; coupling barrels are fixed on a coupling barrel fixing device; the coupling barrels are arranged on the coupling barrel fixing device in an array manner of 4*4 uniform arrangement; and the thermal cycling module device comprises a porous reaction tank for inserting a plurality of test tubes, a reaction tank fixing device, a heating refrigeration sheet, a reaction refrigeration heat radiator and a reaction tank refrigeration heat radiation fan which are integrated form top to bottom. According to the rotary scanning real-time fluorescent quantitative PCR detection system, the developed high-power semiconductor heating refrigeration sheet is adopted to realize temperature circulation of a PCR reaction; and the temperature rising speed is high and the amplification efficiency is high.