2224 results about "DNA extraction" patented technology

A method for isolating nucleic acid which comprises:

• • (a) applying a sample comprising cells containing nucleic acid to a filter, whereby the cells are retained as a retentate and contaminants are removed;
  • (b) lysing the retentate from step (a) while the retentate is retained by the filter to form a cell lysate containing the nucleic acid;
  • (c) filtering the cell lysate with the filter to retain the nucleic acid and remove remaining cell lysate;
  • (d) optionally washing the nucleic acid retained by the filter; and
  • (e) eluting the nucleic acid, wherein the filter composition and dimensions are selected so that the filter is capable of retaining the cells and the nucleic acid.

Additionally, there is provided a substrate for lysing cells and purifying nucleic acid having a matrix and a coating and an integrity maintainer for maintaining the purified nucleic acid. Also provided is a method of purifying nucleic acid by applying a nucleic acid sample to a substrate having an anionic detergent affixed to a matrix, the substrate physically capturing the nucleic acid, bonding the nucleic acid to a substrate and generating a signal when the nucleic acid bonds to the substrate indicating the presence of the nucleic acid. A kit for purifying nucleic acid containing a coated matrix and an integrity maintenance provider for preserving the matrix and purifying nucleic acid is also provided. Further, there is provided a method for quantifying DNA, such as double-stranded or genomic DNA, isolated from cells, such as leukocytes to determine the numbers of leukocytes in a sample of leukoreduced blood.

The invention provides a multifunctional integrated microfluidic nucleic acid analysis chip. The chip is prepared by preparing a combination mould according to 3D printing or micromachining technology, generating a microfluidic substrate according to an injection moulding method, and completing bonding of multiple layers including a chip substrate, a liquid circuit layer, an elastic film, a gas circuit layer and the like by means of plasma cleaning to finally obtain the complete chip. The chip integrates various structures including reagent cavities, mixing cavities, a splitting cavity, a DNA (deoxyribonucleic acid) extraction cavity, amplification reaction cavities, a microvalve, a microchannel and the like and has functions of sample loading, reagent mixing, heating for splitting, DNA adsorption, cleaning, eluting, amplification reaction and the like. The multifunctional integrated microfluidic nucleic acid analysis chip has the advantages that totally-closed automatic operation of 'sample input-result output' is realized, complicated manual operations are avoided, and detection efficiency is improved.

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 discloses a whole blood DNA (deoxyribonucleic acid) extraction kit based on a paramagnetic particle method and an application of the extraction kit. The kit comprises a cell lysis solution, a binding solution, a washing solution I, a washing solution II and a nucleic acid eluent. Compared with kits in the prior art, the kit has the advantages of simple and quick operation, use safety, high extraction efficiency, low manufacturing cost and the like; no toxic reagents or organic solvents are used when the kit is used, a blood sample is not required to be pre-treated, the extraction process can be finished manually and can also be automatically finished in a full-automatic nucleic acid extraction instrument, 1-16 animal tissue samples can be treated each time with a 96-hole reaction plate during automatic extraction, other operation is not required in the extraction process, time is saved for whole blood DNA extraction and detection, and the cost is reduced; DNA in the blood can be quickly extracted with the adoption of the kit, the concentration and the purity of extracted nucleic acid are high, the repeatability is good, and the kit can be used for scientific research and clinical detection.

An integrated gel based microfluidic sample processing device, suitable for forensic investigations at the scene of a crime, including a substrate having a plurality of micro-channels to form at least a DNA extraction chamber in fluidic cooperation with an amplification chamber which in turn is in fluidic cooperation with a separation and detection channel. The micro-channels containing a DNA extraction material and gel based reaction reagents necessary for processing the sample. The device further having electrical contacts for coupling to an external power source and capable of inducing electro-kinetic manipulation of the gel based reagents and DNA extracted from the sample throughout the device.

The invention discloses a listeria monocytogenes nucleic acid chromatography detection kit, its detection method and an application thereof, and belongs to the field of molecular biology and immunology. According to the invention, with the combination of a polymerase chain reaction technology in molecular biology and an immunochromatography technology, rapid detection of listeria monocytogenes in food is accomplished. The kit provided by the invention includes three parts of DNA extraction, PCR amplification and strip detection. The combination of the molecular biology technology and the immunochromatography technology is accomplished by modification of primers and selection of corresponding coatings and markers in strip preparation so as to achieve specific amplification product detection of listeria monocytogenes in food. Compared with a traditional biochemistry method, the method provided by the invention is used to greatly save detection time and simplify operation steps, has characteristics of accuracy, rapidity, convenience and safety, and is suitable for detection of a large number of samples.

The invention discloses a fluorescence quantitative PCR detection kit of hepatitis B virus and an application thereof. The kit is composed of the following independent components: DNA extraction solution I, DNA extraction solution II, DNA extraction solution III, DNA extraction solution IV, positive control interior label, PCR reaction liquid, probe HBV-SP, enzyme mixed liquor containing heat resistant DNA polyase and uracil DNA glycosylase, quantitative hepatitis B virus reference material, hepatitis B virus positive control serum and hepatitis B virus negative control serum, wherein DNA extraction solution I contains 0.2-1.0% of lauryl sodium sulphate (mass/volume), 1.0-4.0% of Triton (volume/volume) and 0.2-1.0mol/L of guanidinium isothiocyanate; DNA extraction solution II contains 100-300mmol/L of 4-HEPES, 100-300mmol/L of sodium chloride with pH of 6.5+/-0.2 and 100-400 mu g/ml of magnetic beads; DNA extraction solution III contains 0.1-1.0% of Triton (volume/volume) and 100-300mmol/L of sodium chloride; DNA extraction solution IV contains mineral oil. The fluorescence quantitative PCR detection kit of hepatitis B virus of the invention can be used for detecting the HBV-DNA concentration in samples of serum, blood plasma or latex and the like.

The invention discloses a method for deleting drug resistant genes of acinetobacter baumannii (AB) through CRISPR-Cas9. The method comprises the steps that firstly, AB is collected clinically, and drug resistance analysis and statistics are conducted after the AB is subjected to drug resistance measurement through a drug sensitive slip method; secondly, the multi-drug resistant AB is subjected to DNA extraction through a lysis-boiling method, and then amplification analysis is conducted by mean of well designed drug resistant gene primers of the AB; and thirdly, according to drug resistant gene detection in the former step, the AB containing an OXA-23 gene is selected, CRISPR/Cas9 and sgRNA plasmids are established and transferred into the AB containing the OXA-23 gene, an OXA-23 gene deletion AB mutant strain is established, and drug resistance analysis is conducted on the OXA-23 gene deletion AB mutant strain. The method is easy to operate and high in knocking-out efficiency, and a novel method and a novel concept are provided for preventing spreading of the drug resistant genes and treating drug resistant bacteria.

The invention discloses a fluorescent quantitative PCR (Polymerase Chain Reaction) detection reagent, a kit and a detection method for an African swine fever virus (ASFV). The detection reagent is targeted to a conserved segment of an ASFV VP72 gene, and comprises a pair of specific primers, a specific probe and an internal labeling probe. The kit comprises a PCR mixed solution, Taq DNA polymerase, DEPC (diethylpyrocarbonate)-H2O, an ASFV positive quality product, an ASFV negative quality product, a quantification standard substance and a pseudovirus internal labeling solution, wherein the PCR mixed solution comprises a pair of specific primers, a specific probe and an internal labeling probe. The detection method comprises the steps of extraction of total DNAs, preparation of reaction components, making of a standard curve by diluting the standard substance, amplifying and result judgment. The reagent and the kit have the advantages of high specificity, high sensitivity, no pollution and high flux, and can be used for accurately detecting infection, inapparent infection or continuous carrying of a toxic host of low-content ASFV. The detection method is quick, specific and sensitive, and can be used for quantitatively detecting a large quantity of samples at the same time.

The invention relates to the microbiology and molecular biology technical field, in particular to a method for extracting pig manure sample total DNA. The method mainly comprises manure sample pretreatment and sample total DNA extraction. Moreover, the method comprises the following steps that: collected fresh manure sample or frozen manure sample is taken as a material; the pretreatment of the manure sample is carried out by means of a PBS buffer solution, a paraformaldehyde solution and anhydrous alcohol so as to obtain manure sample bacteria suspension; and a CTAB solution and beta-mercaptoethanol, etc. are used to carry out cell fragmentation, thereby releasing DNA. The method has the characteristics of simple operation, quickness, high efficiency and ideal repeatability and economical efficiency, etc.; moreover, the extracted DNA has high concentration, purity and yield, and can be directly used in subsequent molecular biology experimental researches such as PCR amplification without purification.

The invention provides a DNA library construction method for prenatal diagnosis. According to the DNA library construction method for the prenatal diagnosis, pregnant woman blood plasma is directly used for library construction, and a DNA extraction step is omitted, so that use amount of the pregnant woman blood plasma can be reduced, at the same time, possibility of deviation of fetal DNA content can be reduced, and cost and time for the DNA library construction are greatly reduced. Another aspect of the invention relates to DNA libraries prepared by any of above preparation methods. The third aspect of the invention relates to a DNA sequencing method. The DNA library construction method has good application prospect.

The invention discloses a method for extracting and purifying spittle DNA, which comprises the following: 1) a step of pre-treatment, which is to contact the spittle with pretreatment lysis solution, remove solid matters adhered onto cells and obtain coarse lysis solution; 2) a step of nucleic acid absorption, which is to mix the coarse lysis solution obtained by the step 1) with lysis binding solution and magnetic bead suspension to form a solution system containing magnetic nano microsphere and DNA composite and is to collect the magnetic nano microsphere and DNA composite from the solutionsystem; and 3) a step of washing, which is to wash the magnetic nano microsphere and DNA composite obtained by the step 2) with washing liquid I and washing liquid II in turn, and dissolve out DNA byusing eluent. When the method provided by the invention is used, the DNA extraction efficiency may reach 90 percent, the extracted DNA can be used in short tandem repeat (STR) composite amplification, DNA sequencing, DNA quantification and other downstream analysis and operation.

The invention belongs to the technical field of food quality and safety detection and particularly relates to a method for rapidly detecting pork, beef, mutton and chicken components in food at the same time through animal genomic DNA (deoxyribonucleic acid) extraction, primer design and UP-M-PCR (universal primers-multiplex-PCR). The four meat components can be distinguished rapidly at the same time according to differential sites of mitochondrial cytochrome b of the animal by using the forward primer sharing and reverse primer specificity strategy and an M-PCR system including five primers. The result shows that the method has the advantages that the detection limit can be up to the pictogram level, the components of the reaction system are free from cross interference and the meat products on market can be detected, verified and distinguished accurately by sampling 80 meat products randomly according to the specificity of the four target meat components. In a word, the invention provides a specific, sensitive and practical multiplex PCR method for rapidly screening four common meat components in food at the same time.

The invention relates to 'a blood DNA preserving card and production method' and belongs to the biology technical field. A blood DNA preserving card comprises fiber-shaped media with the ability of absorbing DNA, and is characterized in that: the salinity is absorbed in the media and contains cell lysing reagent, nuclease inhibitor, a substance allowing the pathogenic bacterium and the virus protein to be denaturalized as well as anti-oxidization inhibitor. These salinities can inhibit the activities of the viruses to prevent the infection, can prevent the nuclease from gradating DNA, so that the blood DNA can be preserved indoor for a long-term. The pure DNA can be obtained from the preserved DNA in the card of the invention by bleaching the salt ions and blood corpuscle leftovers on the card media with the water, thereby avoiding the extraction process, ensuring the operation is simpler, being convenient to be operated in batches and saving the expense of the DNA extraction agent.

The invention discloses an extraction and purification method of total plant endophyte genome DNA for colony analysis. The method comprises the following steps of: shearing and grinding the surface bacteria removed fresh plant tissues into paste, soaking the plant tissues into sterilized phosphate buffer solution, performing thermostatic shaking for 1 hour in a table concentrator to separate out microbial cells from the plant tissues, standing, transferring the suspension to a centrifuge tube, performing centrifugal collection on microbial bacteria, then adding lysing solution, shaking and uniformly mixing the solution to release DNA from the cells, repeatedly extracting protein by chloroform-isoamylol, performing centrifugal sedimentation with isopropanol and washing with 70 percent ice ethanol, performing purification by using a DNA specific centrifugal adsorption column to obtain total crude extracted genome DNA, and finally, performing polymerase chain reaction (PCR) amplificationby using special primers to obtain high-quality plant endophyte specific DNA fragments for subsequent analysis. The invention has the advantages that: the method is simple and effective, high in quality, low in pollution and the like, and provides high-quality guarantee for comprehensively and completely researching a plant endophyte colony structure.

Environmental samples typically include impurities that interfere with PCR amplification and DNA quantitation. Samples of soil, river water, and aerosol were taken from the environment and added to an aqueous buffer (with or without detergent). Cells from the sample are lysed, releasing their DNA into the buffer. After removing insoluble cell components, the remaining soluble DNA-containing extract is treated with N-phenacylthiazolium bromide, which causes rapid precipitation of impurities. Centrifugation provides a supernatant that can be used or diluted for PCR amplification of DNA, or further purified. The method may provide a DNA-containing extract sufficiently pure for PCR amplification within 5–10 minutes.

The present invention relates to a PCR-RFLP identification method for 7 kinds of Chinese south inshore oysters. Said identification method includes the following several main steps: making genome DNA extraction, PCR amplification of object fragment, selecting proper incision enzyme combination, utilizing selected incision enzyme to enzyme-cut the PCR product and utilizing agarose gel electrophoresis to make detection.

The present invention is a portable device for detecting microorganisms in a food sample. The device includes a DNA extraction tube, a chip, and a portable control unit. The DNA extraction tube extracts DNA from the sample in an aqueous solution. The extraction tube includes successive chambers that are configured to mince, filter, and purify DNA in the form of an aqueous eluate. The chip includes reaction chambers disposed for a polymerase chain reaction to amplify a specific DNA fragment in the eluate for detection, and the portable control unit is configured to receive the chip and analyze the amplified DNA on the chip.

The invention relates to a method for obtaining a capsicum phytophthora resistance candidate gene and a molecular marker, and application. The method is used for obtaining the capsicum phytophthora resistance candidate gene by utilizing capsicum phytophthora transcriptome and whole-genome sequencing data information, differentially-expressed gene identification, bioinformatics analysis, molecular marker development and phytophthora inoculation identification and belongs to the technical field of capsicum biology. The method comprises the following steps: sequencing a phytophthora resistant and susceptible gene pool transcriptome obtained after phytophthora inoculation of an F2 population constructed by capsicum highly-resistant and highly-susceptible phytophthora materials, performing expression analysis and functional annotation on differential genes, extracting DNAs (Desoxvribose Nucleic Acid) of a capsicum phytophthora highly-resistant and highly-susceptible phytophthora material genome, performing primer design and PCR (Polymerase Chain Reaction) amplification, performing sequence difference analysis and SNP site identification, performing SNP specific primer design and validity verification, and performing other steps to efficiently obtain the capsicum phytophthora resistance candidate gene and the molecular marker. According to the method, the capsicum phytophthora resistance candidate gene can be accurately identified, and the effective molecular marker can be developed.

The invention discloses a plant endophyte 16S rRNA gene amplification method and application. The amplification method includes the following steps of plant pretreating; plant-sample total DNA extracting; sample 16S rRNA gene amplifying, wherein sample 16S rRNA gene amplifying comprises 16S rRNA gene total-length emulsion PCR amplifying and 16S rRNA gene hypervariable-region/conserved-region amplifying sub-amplifying; high-throughput sequencing and biological information analyzing based on a Illumina platform, wherein high-throughput sequencing and biological information analyzing based on the Illumina platform comprises plant endophyte 16S rRNA gene amplicon purifying and recycling, amplicon sequencing library establishing, Illumina HiSeq sequencing and sequencing data bioinformatics analysis. The gene amplification method is used for high-throughput-sequencing plant disease detection and phytophagous animal enteric microorganism detection. Sequencing analysis of plant endophytic bacteria is carried out with the high-throughput sequencing technology, the data size is larger, the detection result is more complete, pollution of plant hosts is reduced to the maximum degree, the result multiformity is higher, and the cost is low.

The invention discloses a human feces DNA extraction method. The prior art is complicated and time-consuming; and the trace DNAs extracted by the previous sampling method is too little to perform molecular diagnosis. The extraction method uses selective sample collection PSC equipment for feces samples and adopts a low temperature preservation method. The extraction method comprises the following steps: picking a sample by using a sterilized gun head or sterilized blade and directly placing the picked sample on ice; and adding a feces lysis solution, fully cracking and evenly mixing, adding bovine serum albumin, centrifuging to remove the supernatant, adding protease K to perform a digestion reaction at 58 DEG C, then adding a protein precipitation solution to precipitate the protein, centrifuging to obtain the supernatant, adding a silica gel membrane binding solution to fully bind DNAs, centrifuging to obtain precipitates, adding a silica gel membrane washing solution to wash twice, centrifuging, and adding a TE (Tris + EDTA) buffer solution to finally obtain a preservation solution with DNAs. The human feces DNA extraction method has the advantages of high repeatability and stability, simpleness and short time.

An apparatus and method for searching for similar malicious code based on malicious code feature information. The apparatus includes a malicious code registration unit for registering input new malicious code as a new malicious code sample, and extracting and registering detailed information of the new malicious code sample, a malicious code analysis unit for analyzing the detailed information of the new malicious code sample, a malicious code DNA extraction unit for extracting malicious code DNA information including malicious code feature information, a malicious code DNA comparison unit for comparing the extracted malicious code DNA information with malicious code DNA information of prestored malicious code samples, and calculating similarities therebetween, and a similar malicious code search unit for calculating, based on the calculated similarities, all similarities between the new malicious code sample and prestored malicious code samples, and extracting a specific number of malicious code samples.