31 results about "Gene conservation" patented technology

The invention provides a non-lineage animal pairing evaluation method based on genome data, and belongs to the technical field of bioinformatics and protection genetics, the method comprises the following steps: firstly, obtaining whole genome sequencing data of all individuals in a population, and carrying out quality control to obtain a high-quality SNP site set; aiming at all possible male and female pairing combinations in the population, calculating an inter-parent genetic coefficient depKin, an inter-parent heterozygous difference ratio HDR and an inter-parent potential risk load index GRLI, and predicting a fixed proportion Proh of ROH of offspring; performing z-score standardization processing on the four genetic indexes: performing principal component analysis on standardized data, extracting the first two principal components PC1 and PC2, determining a weight according to a variance contribution rate, and calculating a pairing comprehensive score; and generating a pairing candidate recommendation list for each individual according to the comprehensive score, and screening a high-quality pairing scheme. The method disclosed by the invention can be completely independent of pedigree records, and genetic evaluation is directly carried out based on genome data.

The invention discloses a specific primer group, a kit and a method for detecting double photosynthetic bacteria, and relates to the technical field of microbiological detection. The specific primer group is designed based on a conserved region of a photosynthetic reaction center L subunit gene (pufL) of rhodopseudomonas and ectothiorhodospirillum, the conservative property is greater than or equal to 99%, the similarity rate is greater than or equal to 98.5%, specific amplification of target bacteria can be realized, and cross amplification interference is avoided. The matched kit comprises the primer group, 2 * SYBR Green qPCR Master Mix, a positive control component, a negative control component and the like, and the detection method realizes rapid and sensitive detection of the two photosynthetic bacteria by optimizing sample pretreatment, PCR amplification conditions and result interpretation standards. The detection specificity is high, the sensitivity is high (the lowest detection limit reaches 10 < 2 > cfu / mL level), the detection period is short (1-2 hours), the applicability is wide, the method can be used for detecting various samples such as river water, aquaculture water and microbial inoculum fermentation liquor, one set of primers can detect two target bacteria at the same time, the detection cost and the operation complexity are reduced, and the method is suitable for aquaculture environment monitoring and microbial inoculum application quality control.

The application belongs to the technical field of nucleic acid detection methods, and particularly relates to a rapid detection method of avian influenza virus, which comprises using a primer pair and a probe designed for a conserved region of M gene of AIV, and amplifying amplification reagents containing a sample to be detected based on super-fast qPCR technology, wherein the sequence of a forward primer in the primer pair is 5'-GGCCCCCTCAAAGCCGA-3'; the sequence of a reverse primer is 5'-CGTCTACGCTGCAGTCC-3'; and the sequence of the probe is ROX-CACTGGGCACGGTGAGCGT-BHQ2. The method of the application can rapidly and accurately detect whether the avian influenza virus exists in the sample to be detected and the content of the avian influenza virus, and has a good application prospect.

The invention relates to a primer probe combination for identifying E, J and K subgroups of avian leukosis viruses, a detection method and application of the primer probe combination. The detection system for identifying the avian leukosis virus subgroup comprises an upstream primer ALV-E-F, a downstream primer ALV-E-R and a probe ALV-E-P of a gp85 gene conserved region of targeted ALV-E, and an upstream primer ALV-J-F, a downstream primer ALV-J-R and a probe ALV-J-P of an env gene conserved region of targeted ALV-J, the upstream primer ALV-K-F, the downstream primer ALV-K-R and the probe ALV-K-P are used for targeting a gp85 gene conserved region of ALV-K. The detection system for identifying the avian leukosis virus subgroups has no cross reaction with other ALV subgroups and common avian pathogens, and has strong specificity.

The application relates to the technical field of animal virus molecular detection, and particularly discloses a multiple detection method for ASFV, PCV2 and PCV3 suitable for rapid real-time fluorescent PCR. Specific primers and TaqMan probes are designed according to the B646L gene of ASFV and the ORF1 gene conservative regions of PCV2 and PCV3, and the sequences are shown as SEQ ID NO:1-9. The primers and probes are optimized: avoiding strong GC anchoring at the 3' end of the primers, continuous AT series and G base at the 3' end of the probes, and shortening the length of the probes to adapt to rapid real-time fluorescent PCR. The minimum detection limit of the three viruses is not higher than 5 copies per muL, the detection is completed within 30 minutes, there is no cross reaction with 11 common pig pathogenic agents, the coefficient of variation is less than 2%, the sensitivity is high, the specificity is strong, the method is rapid and convenient, and is suitable for rapid diagnosis of mixed infection of ASFV, PCV2 and PCV3.

The invention discloses a dual digital PCR (polymerase chain reaction) detection method for monkey campylobacter jejuni and monkey enterocolitis yersinia, and relates to the technical field of microbiological detection, the detection method comprises the following specific steps: multi-source heterogeneous data collection: collecting monkey feces and intestinal tissue samples, extracting genome DNA (deoxyribonucleic acid) by using a phenol-chloroform method, detecting the concentration and purity of DNA, and detecting the concentration and purity of the DNA; after reaching the standard, storing at-20 DEG C for later use; according to the method, high-sensitivity and specific detection on the two pathogenic bacteria is realized through a combination of well-designed primers and probes, according to the method, a gyrA gene conserved region of monkey campylobacter jejuni CJ and a POL gene conserved region of monkey yersinia enterocolitica YE are used as detection targets, and specific fragments of 249bp and 182bp are amplified respectively; the method effectively avoids interference of non-specific amplification, can simultaneously detect two pathogenic bacteria in a single reaction system through a dual digital PCR technology, accurately calculates target gene copy number concentration through a Poisson distribution formula, and improves detection accuracy and reliability.

The invention belongs to the technical field of molecular biological detection, and particularly relates to an RT-LAMP (Reverse Transcription Loop-Mediated Isothermal Amplification) and immunochromatography combined PEDV (Porcine Epidemic Diarrhea Virus) nucleic acid detection kit and application thereof. According to the kit, the high sensitivity and high specificity of the loop-mediated isothermal amplification technology and the rapid and visual interpretation advantages of the immunochromatography test strip are combined, and on-site, rapid and accurate detection of the PEDV is achieved. The core of the invention comprises a set of specific primer group designed for a PEDV N gene conserved region, an optimized RT-LAMP reaction system and an immunochromatography test strip which is specially treated and is used for detecting an LAMP amplification product. The kit does not need an expensive thermal cycler, the whole detection process is completed within 40 min, the sensitivity can reach 100 copies / [mu] L, and the kit is particularly suitable for rapid diagnosis and epidemic prevention and control of sites such as pig farms and grassroots veterinary stations.

This invention belongs to the field of molecular biology detection technology, specifically relating to a Nipah virus (NiV) nucleic acid detection kit combining RT-LAMP and immunochromatography, and its application. This kit combines the high sensitivity and specificity of loop-mediated isothermal amplification (LAMP) technology with the rapid and visual interpretation advantages of immunochromatographic test strips, enabling on-site, rapid, and accurate detection of NiV. The core components of this invention include: a set of specific primers designed for conserved regions of the NiV gene, an optimized RT-LAMP reaction system, and specially treated immunochromatographic test strips for detecting LAMP amplification products. This kit eliminates the need for reverse transcription of RNA virus nucleic acids and expensive thermal cycling equipment. The entire detection process is completed within 20–30 minutes, with a sensitivity of up to 300 copies / μL. This solution combines the high efficiency of RT-LAMP technology with an intuitive, equipment-free detection method, achieving truly rapid and accurate on-site diagnosis of NiV.

The present application relates to the technical field of plant gene function, and aims to provide a construction method of a homologous silencing system of a Paeonia lactiflora bud dormancy gene, comprising the following steps: designing amplification primers according to gene annotation and a conserved sequence region in a Paeonia lactiflora bud dormancy full-length transcriptome, and constructing a conserved sequence fragment of a target gene into Agrobacterium tumefaciens containing a pTRV vector; taking a rhizome of a treated Paeonia lactiflora annual seedling as an infection material, and placing the infection material in Agrobacterium tumefaciens liquid containing pTRV1, pTRV2 and pTRV2-target gene fragments to perform infection treatment; potting the rhizome and moving the rhizome into a culture room to perform culture, observing phenotype and expression amount differences between the rhizome and a control group in a growth process, and confirming that the homologous silencing system is successfully constructed. The present application first completes miniaturization treatment and batch bacterial liquid infection of an underground rhizome system of Paeonia lactiflora, establishes a virus-mediated homologous function silencing system of a Paeonia lactiflora bud dormancy gene, and realizes the purpose of verifying functions of bud dormancy and even germination and growth related genes.

The invention discloses a sequence combination, a kit and a method for co-detecting a new bunyavirus and an anaplasma phagocytophilum based on RPA-CRISPR, and application of the sequence combination, the kit and the method. Comprising a first RPA primer pair and a first crRNA which are designed according to an SFTSV L segment conserved region, a second RPA primer pair and a second crRNA which are designed according to an AP 16S rRNA gene conserved region, an optimized universal T7 auxiliary primer, a kit prepared by using the sequence combination, and a detection method for detecting the new bunyavirus and the anaplasma phagocytophilum by using the kit. According to the detection method disclosed by the invention, an RPA amplification method and a CRISPR detection method are combined to detect the new bunyavirus and the anaplasma phagocytophilum at the same time, and the bottlenecks in co-amplification efficiency and orthogonal detection sensitivity in the prior art can be effectively overcome.

The invention discloses a primer probe combination and application thereof in simultaneous identification of chlamydia psittaci and chlamydia abortus, and relates to the technical field of animal molecular biology test methods and test reagents. The method comprises the following steps: firstly, designing a universal primer probe for a common conserved region of chlamydia parrot (15 genotypes) and chlamydia abortus ompA genes, then designing an identification primer probe for conserved but different regions of ompA genes of two chlamydia, respectively marking FAM and VIC fluorophores by the two probes, then screening out an optimal primer probe combination and optimal PCR amplification reaction conditions, and finally identifying the ompA genes of the chlamydia parrot (15 genotypes) and the ompA genes of the chlamydia abortus. Finally, the sensitivity, specificity, repeatability and the like of the primer probe combination are verified through experiments. By using the detection method disclosed by the invention, the chlamydia psittaci and the chlamydia abortus can be identified at the same time, all genotypes of chlamydia psittaci can be detected, and the effect is obviously superior to that of the existing known detection method.

The invention discloses a detection system for visually detecting a swine Japanese encephalitis virus based on an RT-RAA-CRISPR / Cas12a system, and the visual detection of the JEV is realized by designing crRNA and RAA amplification primers for a JEV NS1 gene conserved region and triggering the breakage of a double-labeled ssDNA reporter molecule by using the specific recognition and trans-cleavage activity of Cas12a protein to an RAA amplification product. The lower detection limit of the method is 2.04 * 10 < 0 > copies / [mu] L; the clinical sample detection rate is 61.0% and is equivalent to that of 63.4% of PCR (Polymerase Chain Reaction). Therefore, the invention establishes a nucleic acid diagnosis method with strong specificity, high sensitivity, short detection time and simple operation, and the method is also suitable for pig farm on-site epidemic situation screening, animal quarantine ports, grassroots veterinary stations, remote area public health monitoring and other aspects.

The invention is applicable to the technical field of biological detection, and provides a primer probe group, system and method for simultaneously detecting mycoplasma pneumoniae and chlamydia pneumoniae, specific primers and probes are designed aiming at conserved regions of 16S rRNA gene of MP and MOMP gene of CP, the two probes are respectively marked with FAM and ROX fluorophores, and the specific primers and probes can be used for simultaneously detecting mycoplasma pneumoniae and chlamydia pneumoniae. Parallel detection of MP and CP nucleic acids can be realized in a single reaction tube. The detection system provided by the invention comprises the PCR enhancer, so that the detection sensitivity and the amplification efficiency are effectively improved. The method further comprises an optimized nucleic acid extraction step and a reaction procedure, has the advantages of being easy and convenient to operate, rapid in detection, high in sensitivity and high in specificity and is suitable for early-stage rapid diagnosis of clinical respiratory tract samples.

The invention belongs to the field of CRK gene family research, and discloses an identification and expression regulation analysis method of amorphophallus konjac AkCRKs gene family members, which is based on a hidden Markov model of a CRK gene conserved domain, and uses TBtools software to carry out identification and bioinformatics analysis on the AkCRKs gene family members. And the expression characteristics of AkCRKs gene family members in different tissue parts, jasmonic acid, soft rot bacteria (Pcc) and under low temperature stress are analyzed. 19 AkCRKs gene family members are identified in the whole genome of amorphophallus konjac and are non-uniformly distributed on 7 chromosomes. The promoter of the AkCRKs gene family member comprises a plurality of adversity stress response, hormone response, growth and development regulation and other cis-acting elements. The expression of the AkCRKs gene has tissue specificity and participates in response to Pcc, MeJA and low-temperature stress. The research result of the invention provides a basis for further researching the function and adversity stress response mechanism of the amorphophallus konjac AkCRKs gene.

The invention discloses a method and a device for generating a siRNA precursor double-stranded RNA sequence aiming at a target organism in the field of bioinformatics and application of the siRNA precursor double-stranded RNA sequence. The invention aims to solve the technical problem of how to design a precursor double-stranded RNA (Ribonucleic Acid) aiming at specific siRNA (Small Interference Ribonucleic Acid) of a target species. The method comprises the following steps: analyzing a candidate conserved gene set of a target species based on transcriptome and protein data, and performing gene conservative analysis to obtain a target gene of the target species; then intercepting all precursor double-stranded RNA sequences based on transcriptome data of a target gene, and enumerating all available sequence fragments to obtain a candidate double-stranded RNA set 1; and screening from the candidate double-stranded RNA set 1 according to a preset off-target threshold to obtain a candidate double-stranded RNA set 2. The invention can be applied to RNAi prevention and control of pathogens or pests of agricultural related species.

The invention discloses a monkey pox virus triple fluorescence PCR nucleic acid detection kit, a detection method and application. The kit adopts a single-tube triple detection system and comprises (a) a monkey pox virus universal detection primer probe group and a targeted F3L and G2R gene conserved region; (b) a Clade Ib subtype detection primer probe group which targets D14L and A36R gene deletion feature regions; (c) a Clade II subtype detection primer probe group, which is used for targeting J2R and D18L gene feature sequences; and (d) an internal standard detection group which targets the human RNase P gene. By optimizing the concentrations of the primers and probes and four-channel fluorescent labels (FAM universal type, VI-Clade Ib, ROX-Clade II and CY5 internal label), single-tube synchronous detection is realized. The leak detection risk is reduced by adopting a double-target redundancy design, virus screening and subtype identification (Clade Ib / II) can be completed through single detection, and an internal standard system monitors the quality of a sample in the whole process. The kit is suitable for clinical early diagnosis, epidemic prevention and control and strain traceability.

The invention discloses a triple fluorescent RT-PCR (Reverse Transcription-Polymerase Chain Reaction) detection kit and a detection method for simultaneously detecting an avian influenza virus, a duck tembusu virus and a novel duck reovirus, specific primers and TaqMan probes are respectively designed aiming at gene conserved regions of the three viruses, reaction conditions are optimized, and the specific primers and TaqMan probes can be used for simultaneously detecting the avian influenza virus, the duck tembusu virus and the novel duck reovirus. A TaqMan triple real-time fluorescent quantitative RT-PCR diagnosis method capable of simultaneously detecting three viruses is established and is applied to clinical sample detection. Results show that the established triple fluorescence RT-PCR method has high sensitivity and strong specificity, can meet the requirements of high throughput and rapid detection, and has high application value.

The invention discloses a qPCR (quantitative polymerase chain reaction) method for quantitatively analyzing human cells in a non-humanized animal model, which comprises the following steps: taking a non-humanized animal model sample, and extracting genome DNA (deoxyribonucleic acid); the method comprises the following steps: designing and synthesizing a specific primer probe combination aiming at a human COX1 gene conserved sequence; the method comprises the following steps: cloning a specific fragment of a human COX1 gene to a plasmid vector, and preparing a linearized plasmid standard substance; carrying out 10-fold gradient dilution on the linearized plasmid standard substance to obtain a standard substance gradient solution, and fitting a linear regression curve; taking the extracted DNA as a template, adding a primer probe combination and a qPCR premixed solution, and carrying out qPCR amplification; and calculating the human cell content in the sample according to the linear regression curve. The method provided by the invention has the advantages of stable standard substance, strong species specificity, high detection sensitivity, cross-species universality and simple operation, can effectively avoid animal matrix DNA interference, solves the problems of quantitative deviation and low-abundance cell detection, and reduces the detection cost.

The invention relates to the technical field of biology, in particular to a primer probe group for detecting avian reovirus and serum type 4 avian adenovirus and application of the primer probe group. According to the invention, an ARV / FAdV-4 double ddPCR detection system and method are developed for the first time on the basis of an S1 gene of an ARV S1133 strain and a hexon gene conserved region of FAdV-4GX2019-010. A sensitivity test shows that the lowest detection limits of ARV and FAdV-4 are 32.67 copies / mu L and 5.27 copies / mu L respectively. Specificity experiments show that the specificity reaches 100%. Clinical sample detection shows that the result is completely consistent with that of qPCR. Therefore, the detection method provided by the invention has the characteristics of high sensitivity, good repeatability, strong specificity and the like, and provides an effective detection technical means for absolute quantitative detection of ARV and FAdV-4.

The invention discloses an avian leukosis virus subgroup rapid detection kit and a detection method thereof, and belongs to the technical field of animal inspection and quarantine. Aiming at the defects of the existing detection technology, a conserved sequence of an env gene shared by A, B and K subgroups and a conserved sequence of a specific env gene of a J subgroup are selected as target regions, specific primer probes are designed, a reaction system and an amplification program are optimized, and the kit is assembled. The kit adopts a fluorescent RT-PCR (Reverse Transcription-Polymerase Chain Reaction) method, can simultaneously detect A, B, J and K subgroups and accurately distinguish endogenous and exogenous viruses without cross interference. The kit has the characteristics of high detection speed, high sensitivity, strong specificity, simplicity and convenience in operation and the like, is suitable for detecting various samples such as chicken plasma, seminal fluid and cloaca swab, can improve the breeding chicken purification efficiency, shorten the period and reduce the economic loss of the poultry industry, and has remarkable application value.

The invention provides an RNA (Ribonucleic Acid) molecular marker for judging early gonad differentiation of young river snails in China and an identification method. Dmrt1 and Foxl2 genes are important genes related to gonad differentiation of river snails. The method comprises the following steps: designing and verifying primers SEQ ID NO: 1-2 according to conserved regions of Dmrt1 and Foxl2 genes; the method comprises the following steps: collecting gonad tissue samples in a young snail stage and a sexual maturity stage, wherein n is greater than or equal to 120, extracting total RNA (Ribonucleic Acid) of a field snail tissue, carrying out reverse transcription to obtain cDNA (Complementary Deoxyribonucleic Acid), detecting a double-gene expression quantity through qRT-PCR (Quantitative Reverse Transcription-Polymerase Chain Reaction), calculating a standardized expression value by taking beta-actin as an internal reference, and establishing a discrimination function D = log2 (Dmrt1 / Foxl2); 1, judging that the juvenile snails of the field snails start gonad differentiation; according to the invention, the problems of deficiency of specific molecular markers for sex differentiation of the young escargots in China and indefinite gonad differentiation time point are solved in a breakthrough manner, accurate locking of the gonad differentiation time of the young escargots is realized, the gonad differentiation time is 15-20 days earlier than morphological differentiation, and verification of more than one hundred of samples shows that the identification accuracy reaches 100%.

The invention relates to the technical field of biological detection, and discloses a primer, a probe and a kit for detecting aeromonas hydrophila based on fluorescent PCR (Polymerase Chain Reaction) and application of the primer, the probe and the kit. A primer and fluorescent probe combination is obtained by designing and screening a specific gene conserved region of aeromonas hydrophila. The specific amplification of the aeromonas hydrophila gene based on the fluorescent PCR technology can be successfully realized. According to the method, the detection accuracy and sensitivity are improved, the aeromonas hydrophila can be detected in the early stage of bacterial septicemia of silver carps and bighead carps, prevention and control of bacterial septicemia of silver carps and bighead carps are facilitated, and the basic requirements of field detection of farms can be met.

The invention discloses a primer group for simultaneously detecting porcine acute diarrhea syndrome coronavirus and bat HKU2 coronavirus, and belongs to the technical field of biology. The primer combination disclosed by the invention can be used for simultaneously detecting the porcine acute diarrhea syndrome coronavirus (SADS-CoV) and the bat HKU2 coronavirus, and the lowest limit of detection (LOD) is 20 copies per reaction; the primer provided by the invention is combined with three gene conserved regions of SADS-CoV and HKU2 coronavirus, and has relatively high specificity. The method has high reference value for nucleic acid diagnosis of SADS-CoV and HKU2 coronavirus, and is suitable for popularization and application.

This invention discloses a qPCR primer-probe combination, kit, and method for specifically detecting 16SrII group areca etiolated phytoplasma, belonging to the interdisciplinary field of molecular biology and plant pathology. The primer-probe combination includes a forward primer qTuf-F4-2 (SEQ ID NO:1), a reverse primer qTuf-R4 (SEQ ID NO:2), and a fluorescent probe qTuf-P4-1 (SEQ ID NO:3), targeting 16SrII group phytoplasma. tuf This gene conserved region demonstrates high specificity for 16SrII group areca yellowing phytoplasma in qPCR reactions, exhibiting no cross-reactivity with 16SrI group, 16SrXXXII group phytoplasma, areca genome, common pathogens, or endophytes. The limit of detection is 14.35 copies / μL, and the sensitivity meets the requirements for detecting low-load samples. The coefficient of variation for intra- and inter-group repeatability is less than 1%, indicating good stability. The kit and detection method developed based on this combination are simple to operate, highly sensitive, and provide stable results, making them suitable for the accurate diagnosis, pathogen typing, and field epidemic monitoring of areca yellowing disease, providing key technical support for the scientific control of 16SrII group phytoplasma.

The invention belongs to the technical field of biology, particularly relates to a primer probe combination, a kit and a detection method for rapidly and simultaneously detecting bovine astrovirus and bovine norovirus, and aims to solve the problems of long detection time and complex detection equipment in the existing method for simultaneously detecting bovine astrovirus and bovine norovirus. According to BAstV and BNoV gene sequences disclosed by GenBank, the specific primer group for detecting the bovine norovirus and the bovine astrovirus is designed for BAstV ORF2 and BNoV RdRp gene conserved regions by adopting Oligo 6.0 software, and the primer pair, the probe and the method have the advantages of simplicity and convenience in operation, high specificity, high detection rate, high sensitivity, good repeatability and the like.

The application relates to the field of biotechnology, and in particular to a monkeypox virus typing reagent, a kit and application thereof. A set of primers of the monkeypox virus, a West African type and a Central African type shared probe of the monkeypox virus, and a probe of the Congo Basin (Central Africa) branch type are designed according to the conservative regions of J1L genes or J3R genes of the monkeypox virus.

This invention discloses a sequence combination, kit, method, and application for the co-detection of novel Bunyavirus and Anaplasma phagocytophila based on RPA-CRISPR. It includes a first RPA primer pair and a first crRNA designed based on the conserved region of the SFTSV L fragment, a second RPA primer pair and a second crRNA designed based on the conserved region of the AP 16S rRNA gene, an optimized universal T7 helper primer, a kit prepared using this sequence combination, and a detection method for novel Bunyavirus and Anaplasma phagocytophila using this kit. The detection method of this invention combines RPA amplification with CRISPR detection to simultaneously detect novel Bunyavirus and Anaplasma phagocytophila, effectively overcoming the bottlenecks of existing technologies in co-amplification efficiency and orthogonal detection sensitivity.

The present invention relates to the field of aquatic pathogen detection, specifically a fluorescent quantitative RT-PCR detection kit and detection method for screening blue crab reovirus-free crab breeders, wherein the kit detects the ORF region of the blue crab reovirus genome fragment VP11. The detection primers are designed based on the conserved region of the gene in the current epidemic strain and the early strain, avoiding the gene mutation site and ensuring the universality of the detection method. The sensitivity of the kit of the present invention is significantly higher than the currently reported blue crab reovirus detection method, and it has strong specificity. It has no specific amplification for 5 common crustacean pathogens, reduces the detection cost, improves the detection efficiency, and can meet the needs of screening blue crab reovirus-free crab breeders under minimally invasive conditions.

The invention relates to a respiratory syncytial virus nucleic acid detection primer and a kit, and belongs to the technical field of biological detection. The invention aims to solve the technical problems of tedious operation, easy pollution, low sensitivity, easy influence by variants, high false negative risk and the like in the existing RSV detection method. The invention provides an RSV-containing PCR (Polymerase Chain Reaction) detection primer group and a kit, the RSV-containing PCR detection primer group comprises a specific primer and a probe designed aiming at an RSV matrix protein M gene conserved region, and an internal standard primer probe, and the concentration of each component and a reaction system are optimized. The kit has high specificity and sensitivity, RSV RNA as low as 200 copies / mL can be detected, automatic detection is adapted, aerosol pollution and false negative results are effectively avoided, the detection sensitivity to clinical samples reaches 97.12%, the specificity reaches 99.53%, and the detection accuracy and efficiency are remarkably improved.

The invention relates to the technical field of nucleic acid detection, in particular to a CRISPR (clustered regularly interspaced short palindromic repeats) one-step nucleic acid detection system and application, the detection system comprises an isothermal amplification system and a CRISPR reaction system, the isothermal amplification system comprises isothermal amplification primers aiming at a target gene conserved region and nuclease-free water, and the CRISPR reaction system comprises a Cas protein, PAM-free sgRNA or PAM-free crRNA, a fluorescent reporter probe and nuclease-free water. Due to the adoption of the design of PAM-free sgRNA or PAM-free crRNA, the activity of Cas enzyme is limited in the first amplification stage, the CRISPR system cannot cut a detected template and an amplified template in the amplification stage, and the CRISPR system is activated through temperature regulation and control in the second stage to carry out nucleic acid detection, so that the detection sensitivity is remarkably improved.