32 results about "Analysis pcr" patented technology

Disclosed are genomic sequences for nine strains of Cronobacter spp. (C. sakazakii—696, 701, 680; C. malonaticus—507, 681; C. turicensis—564; C. muytjensii—530; C. dublinensis—582; C. genomosp1—581) and compositions, methods, and kits for detecting, identifying and distinguishing Cronobacter spp. strains from each other and from non-Cronobacter spp. strains. Some embodiments describe isolated nucleic acid compositions unique to certain Cronobacter strains as well as compositions that are specific to all Cronobacter spp. Primer and probe compositions and methods of use of primers and probes are also provided. Kits for identification of Cronobacter spp. are also described. Some embodiments relate to computer software methods for setting a control based threshold for analysis of PCR data.

The invention discloses a construction method of a date tree SSR (Simple Sequence repeat) marker molecular genetic map. The construction method comprises the following steps of: (1) extracting genomic DNAs (deoxyribonucleic acids) of parents and offsprings of a date tree to be detected; (2) screening SSR primers by using parent materials; (3) carrying out PCR (Polymerase Chain Reaction) amplification by respectively using the SSR primers and respectively taking the genomic DNAs (deoxyribonucleic acids) of the parents and offsprings of the date tree to be detected as templates; and (4) analyzing PCR amplification results by using biology software, and constructing the date tree SSR marker molecular genetic map. According to the construction method, the high-efficiency low-cost date tree SSR marker technical system is constructed, and obtained SSR markers and a core primer group have the advantages of high efficiency, stability, co-dominance and the like. The set of markers can be used for researches such as the date tree genetic map construction, genetic diversity analysis, identification of variety, molecular mark assisted breeding and the like and promoting the development of date tree genetics and breeding technique.

The invention provides a method for carrying out serotyping on streptococcus pneumoniae by using MLPA (multiplex ligation dependent probe amplification). The method comprises the following steps of: firstly, extracting a sample genome DNA of streptococcus pneumoniae to be detected; carrying out pre-amplification on the extracted genome DNA by using a PCR pre-amplification primer; carrying out hybridization on pre-amplification products of the genome DNA by using an MLPA probe; adding a corresponding fluorescence detecting probe in a reaction system; connecting hybridized MLPA probes by using ligase, and carrying out PCR amplification on the hybridized and connected MLPA probes by using an MLPA universal amplification primer; and analyzing PCR products by using a multicolor fluorescence dissolution curve, so that the serotyping on streptococcus pneumoniae can be realized. The detecting method disclosed by the invention is good in specificity and high in sensitivity; according to the method, 10 kinds of serotypes can be subjected to genotyping simultaneously just in 2-4 hours, so that multiple uncapping operations in traditional MLPA detection are avoided, and the pollution possibility is reduced, therefore, the method can meet high-throughput sample detection, and is especially applicable to inspection departments.

Disclosed are diagnostic methods for determining a subtype of methicillin-resistant Staphylococcus aureus (MRSA) in a biological sample of a mammal. Methods include providing a biological sample of the mammal, performing a PCR analysis of the biological sample, and analyzing the PCR amplicons with respect to their sizes so as to determine for type I, type II, type III, type IV or type V MRSA that may be present in the biological sample. Further example embodiments include using at least one mecA primer pair and/or using at least one Staphylococcus aureus nuc primer pair in the PCR analysis. Further disclosed are methods for screening populations for MRSA, and methods of treating a mammal testing positive for Type IV MRSA. Also disclosed are kits for determining a MRSA subtype in a mammal and isolated primers that may be used in the present methods and kits.

The invention belongs to a molecule detection method of Fusarium graminearum to medium resistance level bacterial strain of carbendazim. The method can be used for monitoring drug resistance of Fusarium graminearum to carbendazim and prevalence warning, wherein Fusarium graminearum can cause wheat scab. The detection method includes the following three main steps: (1) nuclear genome DNA of bacterial strain to be detected is respectively extracted: (2) internal and external primer pairs are utilized to carry out nested PCR, thus obtaining a target segment; (3) the target segment is respectively restricted by restriction enzymes HindIII and TaaI (Tsp4CI), and genotype of medium resistance level bacterial strain can be accurately identified by virtue of a restriction product electrophoresis spectrum. By adopting PIRA-PCR (primer-introduced restriction analysis PCR) technology, quantity of Fusarium graminearum with medium drug resistance in field and ratio thereof in group can be rapidly and accurately detected. Detection accuracy reaches more than 95%.

The invention provides a polymerase chain reaction (PCR) detection method of hepatitis B virus (HBV) genotyping. The method comprises the following steps: a primer is designed according to the entire genome sequence of the HBV genotype; in the existence of the primer, DNAs are extracted from a serum sample to perform PCR amplification in a fluorescence PCR meter, the PCR product is analyzed according to the melting curve, the genotype is judged according to the Tm value of the PCR product and the HBV genotyping can be realized. The PCR melting curve method used in the invention is convenient to operate, the detection result of the method has higher coincidence rate with the detection result obtained by adopting a commercial genotyping kit; and the method can be used in the clinical detection of the HBV genotype. On the basis of the PCR technology, the PCR melting curve method which is to judge the HBV genotype according to the Tm value, is provided in the PCR detection method, thus the HBV genotype can be identified conveniently, rapidly and accurately.

The invention provides a method for fast detecting genome DNA residues in a total RNA sample. The method comprises the steps that 1, according to genetic information of a to-be-detected biological material, a gene containing one or more intron sequence is selected as a target, and for the intron sequence, PCR primers are designed; 2, the total RNA of the to-be-detected biological material is extracted, the total RNA or cDNA obtained through reverse transcription of the total RNA is used as a template, and the primers in the step 1 are used for PCR amplification; 3, a PCR amplification productis analyzed. According to the method, only one pair of the primers are designed, so that whether or not the genome DNA residues exist in the total RNA and cDNA samples of the biological material can be detected simply, conveniently and fast; the method provides support for the accuracy of gene expression analysis and has important scientific significance for promoting fundamental research of the molecular biology.

The invention discloses a method for identifying the varieties of low-phenol cotton. The genome DNA of each tested material is extracted; three SSR (simple sequence repeats) primers are respectively selected on 26 linkage groups of the cotton, so that the genome DNA of each tested material is used as a template for PCR (polymerase chain reaction) amplification; the PCR amplification result is analyzed; a fingerprint atlas of each variety of the low-phenol cotton is built; different varieties of feature primers are screened out; 53 pairs of primers express stable polymorphism among different tested materials; at least two pairs of feature primers are used and combined, and the varieties of the low-phenol cotton can be distinguished; the powerful technical support is provided for the identification, the development and the protection of the low-phenol cotton in future.

The present invention discloses a primer set for detecting bovine nornovirus and bovine coronavirus, the primer set comprises a primer for detecting the bovine nornovirus and a primer for detecting the bovine coronavirus, and the nucleotide sequences of the primer set are as shown in SEQ ID NO. 1-SEQ ID NO. 4. The invention also discloses a double RT-PCR method for detecting the bovine norovirus and the bovine coronavirus, and the steps are as follows: (1) synthesizing the primer set for detecting the bovine norovirus and the bovine coronavirus; (2) extracting RNA in a to-be-tested sample, andperforming inverse transcription on the extracted RNA as a template to obtain cDNA; (3) performing PCR amplification reaction on the cDNA obtained in the step (2) as a template by use of the primer set synthesized in the step (1); and (4) analyzing a PCR amplification product. The double RT-PCR method is simple, rapid, economical and efficient, and can greatly save detection time and improve detection efficiency.

The invention discloses an EST-SSR molecular identification method for E.wushanense and an easily mixed species thereof, and belongs to molecular markers and medicinal material identification in the technical field of molecular biology. An EST-SSR primer group for identification of E.wushanense and the easily mixed species thereof include 10 pairs of EST-SSR core primers and common M13 primers having the sequences of SEQ ID NO:1, 2...20 and 21. The identification method includes the steps: (1) extracting total DNA of a sample by a CTAB method; (2) amplifying by PCR by using total DNA of the sample to be determined as a template; (3) analyzing a product of PCR amplification; and (4) constructing a phylogenetic tree. An application comprises identification of E.wushanense and the easily mixed species thereof. The 10 SSR molecular markers provided by the invention have good polymorphism, high specificity and stable amplification results, can rapidly and accurately identify E.wushanense and the easily mixed species thereof, provide technical support for promotion of the quality standard of epimedium medicinal materials, and are beneficial to development and utilization of the epimediummedicinal materials.

The invention discloses a method for analyzing haplotype of a PCR product employing non-synchronous synthesis sequencing of two nucleotides. Through real-time synthesis sequencing of two nucleotides, two different nucleotides are simultaneously added for each sequencing reaction; the synthesis sequencing of different DNA templates in the PCR product is not synchronous, so that information of different base groups on two adjacent SNP sites is provided by different sequencing reactions; and association analysis is carried out according to sequencing information on different SNP sites obtained by these non-synchronous synthesis sequencing reactions, so as to determine the haplotype of the PCR product. The analysis range is expanded; the operation is simple; the analysis cost can be reduced; the method is suitable for haplotype analysis of a plurality of mixing samples; the ratio of individual haplotypes in the mixing samples can be directly determined; and ucleic acid markers can be searched and screened from large samples.

The invention provides specific SS-COI (species-specific-COI) primers for Frankliniella intonsa, a kit containing the primers and used for specifically detecting Frankliniella intonsa, an applicationof the primers or the kit containing the primers to specific detection of the Frankliniella intonsa and a specific SS-COI primer based PCR rapid detection method for the Frankliniella intonsa. The SS-COI primers comprise Fi63-4F:5'-CGCTAAGTATTTTAATTCGG-3'(SEQID No.1) and Fi63-4R:5'-AGTGGCACTAGTCAGTTTCC-3'(SEQ ID No.2). The specific SS-COI primer based PCR rapid detection method comprises the following steps: 1) extracting DNA of a sample; 2) performing a PCR amplification reaction with DNA extracted in the step 1) as a template as well as the primers; 3) analyzing a PCR product.

The invention discloses a method, a kit and application for detecting genotypes of two STR locuses of a hepatitis B virus infected related IL-10 gene. The method is STRs-PCR, which comprises the steps of preparation of a template DNA, primer design of two STR locuses, assembly of a PCR amplification system, PCR circulation reaction, capillary electrophoresis, fluorescence detection and analysis of a PCR product. Compared with the prior art, the invention ensures that the two STR locuses of the IL-10 gene are related with the susceptibility of hepatitis B virus infection, determines IL-10.G and IL-10.R as novel related genes of the hepatitis B virus infection, detects the IL-10.G and the IL-10.R of the IL-10 genes by adopting the STRs-PCR method, thereby determining the susceptibility of individuals to the hepatitis B, and providing new genetic counseling contents for the prevention and control of hepatitis B virus infected groups.

The invention relates to a reagent system and kit for NDRG4 gene methylation detection and application thereof. The reagent system or the kit comprises specific primers SEQ ID NO.1 and SEQ NO.2 and a specific probe SEQ ID NO.3, and a report group and a quenching group are connected to the probe. The reagent system further comprises a positive standard substance, a negative standard substance, a buffer solution, dNTPs and DNA polymerase. A use method of the kit comprises the steps that 1, methylation treatment is conducted on a sample DNA; 2, a fluorescent quantitative PCR reaction is conducted on the sample DNA obtained after methylation treatment in the step 1 by means of the specific primers SEQ ID NO.1 and SEQ NO.2 and the specific probe SEQ ID NO.3, a fluorescent quantitative PCR reaction is conducted on the positive standard substance and the negative standard substance simultaneously, and the methylation state of a gene is judged by analyzing fluorescence alteration of the PCR process. The preparation method of the kit is simple, the kit is convenient to use and high in speed, and the detection result is accurate, visual and high in sensitivity.

The invention discloses a kit for detecting bovine circovirus, which comprises a primer for detecting bovine circovirus, an RNA extracting reagent, a reverse transcription reagent, a positive controland a negative control, wherein the nucleotide sequence of the primer is an upstream primer P1: 5 '-GGTGCCGTTGTTGTGTCA-3 ', downstream prim P2: 5'-CAGCCCTGAGTTGCCTTATC-3'; Positive control is BCoV, BNoV, BAstV, BRV, BVDV and BKV, negative control is sterile double distilled water. The invention also discloses an RT-A PCR method comprises the following steps: (1) synthesizing the primers for detecting bovine circovirus; (2) extracting RNA from the sample to be tested, and reverse transcribing the extracted RNA as a template to obtain cDNA; (3) PCR amplification reaction is carried out by usingthe cDNA prepared in the step (2) as a template and the primer set synthesized in the step (1); (4) PCR amplification products are analyzed. Simple, practical, economical and efficient, it can greatlyimprove the detection efficiency and save the detection time.

The invention discloses a method for quickly detecting specific PCR (polymerase chain reaction) products on the basis of analysis on a PCR byproduct pyrophosphoric acid and application of the method.The method includes steps of extracting sample genomes for PCR; (2), designing at least one PCR primer and substituting alpha-sulfo-deoxyadenosine triphosphate for deoxyadenosine triphosphate and carrying out PCR amplification; (3), analyzing the pyrophosphoric acid in PCR amplification products; (4), judging whether the PCR is carried out or not according to analysis results and detecting the content of to-be-detected DNA (deoxyribonucleic acid) in samples. The method and the application have the advantages that whether the samples contain sample DNA or not and the content of the sample DNA can be accurately detected by the aid of the method, and the detection limit of the method can reach a plurality of copies; the method for detecting and analyzing the specific PCR products is high in sensitivity, the specific PCR products can be quickly detected by the aid of the method, operation processes are simple and are easy to implement, and large sample specific PCR products can be quicklydetected by the aid of the method; the method can be used for quickly detecting microorganisms in the industry of food, sanitation and the like and also can be used for analyzing other nucleotide sequences with specific PCR.

The invention relates to a genetic constitution detecting method for a cloned specific T lymphocyte TCR BV CDR3 gene, which comprises the following steps: a single nucleus cell of peripheral blood or a general RNA in tissue sample is extracted and reverse-transcribed to cDNA. The upper reach primer of the TCR BV twenty four families and a common lower reach primer BC are designed according to TCRBV gene; the cDNA is expanded using real-time fluorescent quantification PCR with dye method. The PCR product is analyzed with the melting curve; the negative primary derivative (-dF/dT) of fluorescence intensity change of the PCR product is graphed vs. the temperature (Tm); the peak graph of BV each family is obtained; the BV family PCR product showing single peak is purified and processed with serial analysis; the sequencing results are compared with the TCR Beta chain standard gene sequence. The distribution of the cloned specific T lymphocyte in tissue or peripheral blood is judged according to the appearing situation of single peak in BV each family peak graph; the genetic constitution of the cloned specific TCR BV CDR3 is determined combining the sequencing results. The genetic constitution detecting method for the cloned specific T lymphocyte TCR BV CDR3 gene has important significance in evaluating diseases related to specific cloned T cell.