CN104911279A - Loop-mediated isothermal amplification kit for detecting newcastle disease virus and application method - Google Patents
Loop-mediated isothermal amplification kit for detecting newcastle disease virus and application method Download PDFInfo
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- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/70—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving virus or bacteriophage
- C12Q1/701—Specific hybridization probes
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6844—Nucleic acid amplification reactions
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
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Abstract
The invention discloses a loop-mediated isothermal amplification kit for detecting newcastle disease virus and an application method thereof. The kit is characterized by comprising an HN gene loop-mediated isothermal amplification primer mixing solution, an loop-mediated isothermal amplification reaction premixing solution, a newcastle disease virus positive sample and a negative quality control product, wherein the primer set is characterized by being designed through newcastle disease virus HN gene conserved domain; the primer set comprises a pair of inner primers (FIP and BIP), a pair of outer primers (F3 and B3) and a pair of loop primers (LF and LB) and can be used to the direct visible fluorescence method, the UV irradiating visible fluorescence method, the gel electrophoresis method, the real-time fluorescence detection method, the real-time turbidity detection method and the micro-total analysis method. The application method of the kit is characterized in that an isothermal amplification fluorescence detection system can be adopted for real-time quick detection of the newcastle disease virus. According to the loop-mediated isothermal amplification kit, the technical means for quickly detecting the newcastle disease virus of the livestock are provided, and high in specificity and sensitivity.
Description
Technical field
The present invention relates to a set of primer sets for viral rapid detection, test kit and using method, particularly relating to a set of loop-mediated isothermal amplification (LAMP) primer group, test kit and using method thereof for detecting Avian pneumo-encephalitis virus, belonging to Avian pneumo-encephalitis virus detection field.
Background technology
Newcastle disease (Newcastle disease, ND), also known as philippine fowl disease, pseudo-checken pest, be a kind of by Avian pneumo-encephalitis virus (Newcastle disease virus, NDV) cause with respiratory tract and the hemorrhage deadly infectious disease for characteristic pathology of alimentary canal mucous membrane.ND has high degree in contact infectivity, and velocity of propagation is exceedingly fast, and during acute onset, case fatality rate is close to 100%, serious harm poultry farming, therefore OIE (OIE) is classified as one of zoonosis that must report.
Avian pneumo-encephalitis virus belongs to paramyxovirus section (Paramyxoviridae) paramyxovirus subfamily (Paramyxovirinae) fowl Rubulavirus (Avulavirus), cyst membrane is had outside virus particle, cyst membrane surface has fine prominent, and its genome is a sub-thread strand RNA being about 15kb.The genome non-segmented negative of NDV, 6 kinds of structural protein of encoding: nucleocapsid protein (NP), phosphorylated protein (P), stromatin (M), fusion rotein (F), hemagglutinin-neuraminidase albumen (HN) and there is the dependent RNA polymerase albumen (L) of RNA.P, NP and L are called inner albumen, form ribosome (RNP) with viral RNA; M, F and HN are then called outside albumen.
HN albumen is that a kind of existing hemagglutinin (HA) is active, has again the glycoprotein that neuraminidase (NA) is active, is the important virulence protein of NDV and protective antigen.HA activity can make viruses adsorption on the acceptor of corresponding target cell, and NA activity then destroys cell receptor, from cells infected surface releasing virus particle.HN albumen can also promote that film merges simultaneously, attracts F protein fully close to this site, then starts virus and cell membrane fusion.5 ' the non-coding region (UTR) of HN protein mRNA is one of factor of influence of NDV virulence, if lacked, NDV copies and can not be affected, but can reduce the virulence of virus.In addition, if HN albumen the 526th amino acids is undergone mutation, the activity of HN albumen will weaken, and copying of NDV will affect by it, and virulence reduces.There is extremely important effect 5 amino acid sites (the 1st, 2,3,4,6) of HN protein N terminal intracellular region location to the fusion of virus and cell, virus virulence and HN albumen and M albumen.This series of studies shows, the tissue tropism of virus depends on HN albumen, and HN is the necessary factor of NDV virulence, and the virulence that jointly decide NDV with F gene is strong and weak.
Ring mediated isothermal amplification (LAMP) method is a kind of new nucleic acid amplification technologies invented by Japanese doctor Notomi in 2000.Its ultimate principle is utilize the primer of 4-6 particular design and have the archaeal dna polymerase of strand-displacement activity, increases under constant temperature to target sequence.RT-LAMP technology have high specificity, susceptibility high, be swift in response, the feature such as equipment requirements is low.The method is through the development of more than ten years, obtain constantly perfect, especially in result judgement, method and the real-time fluorescence detection method of existing multiple macroscopic are used widely, can control from source and be uncapped the Aerosol Pollution caused by electrophoresis, be particularly suitable for field quick detection and bed is other detects.The present invention establishes a set of NDV HN chimeric gene loop-mediated isothermal amplification kit and detection method, for the detection of poultry Avian pneumo-encephalitis virus and investigation and analysis provide a kind of easy, approach fast newly.
Summary of the invention
Main purpose of the present invention is to provide a set of loop-mediated isothermal amplification (LAMP) primer group for NDV HN chimeric gene rapid detection, is made up of HN gene loop-mediated isothermal amplification (LAMP) primer group, can be used for the loop-mediated isothermal amplification detecting Avian pneumo-encephalitis virus.
Described NDV HN chimeric gene loop-mediated isothermal amplification (LAMP) primer group is the HN gene nucleic acid sequence of the different subtype Avian pneumo-encephalitis virus by downloading in a large amount of comparison Genbank, designed by conserved regions fragment sequence.Wherein, HN gene primer group comprises a pair outer primer, a pair inner primer and a pair ring primer, and inner primer and outer primer become by the proportions of 6: 1-12: 1, and preferred proportion is 8: 1, and the ratio of ring primer and inner primer is 1: 2.
Described loop-mediated isothermal amplification premixed liquid can be make amplified production send a kind of commercial kit IsothermalMaster Mix of fluorescent signal or other test kits of equivalence, by Bst polysaccharase, dNTP, trimethyl-glycine, MgSO
4with buffered soln composition, and reversed transcriptive enzyme other reversed transcriptive enzymes as AMV reverse transcriptase or equivalence are formed.Also introduce one group of Avian pneumo-encephalitis virus positive quality control product and negative quality control product in test kit simultaneously.Described Avian pneumo-encephalitis virus positive quality control product can be the total serum IgE extracted by the recombinant plasmid of Avian pneumo-encephalitis virus synthetic fragment structure, in-vitro transcription RNA or Avian pneumo-encephalitis virus LaSota strain; Described negative quality control product is the medium not containing newcastle disease virus gene pack section.
The cumulative volume of described NDV HN chimeric gene loop-mediated isothermal amplification system is 25 μ L, comprising HN gene loop-mediated isothermal amplification (LAMP) primer group solution 6 μ L, loop-mediated isothermal amplification premixed liquid 14 μ L, sample to be tested or each 5 μ L of negative quality control product; The amplification program of ring mediated isothermal amplification fluorescence detecting system is: 1. 65 DEG C, 30min, 2. 98 DEG C-80 DEG C, 0.05 DEG C/s, 10min.
Described NDV HN chimeric gene detection kit, in real-time fluorescence detection method, each reaction fluorescent signal reaches between proliferation time corresponding to maximum value and the logarithmic value of Avian pneumo-encephalitis virus template ribonucleic acid extension rate, in thinning ratio 1: 10-1: 100000 scope, there is linear relationship, R
2> 0.995; The detection limit of the method HN gene can reach 1: 100000 template ribonucleic acid.
Analysis and the decision method of amplification comprise:
1) the amplification curve platform fluorescent value that records of ring mediated isothermal amplification fluorescence detecting system is higher than background noise signal 5 times and the judgement that solubility curve is single sharp peak is positive findings, amplification curve platform fluorescent value is negative findings lower than the judgement of background noise signal 5 times, and amplification curve platform fluorescent value is higher than background noise signal 5 times but the judgement that solubility curve is not single sharp peak is nonspecific cross interference signal.
2) when sample to be tested HN gene loop-mediated isothermal amplification pipe is all negative findings, then Avian pneumo-encephalitis virus is not detected; Sample to be tested is positive findings, then have newcastle disease virus infection; The amplification curve of HN gene with reading during 15min for benchmark.
Accompanying drawing explanation
Fig. 1 chick embryo allantoic liquid Hemagglutination titer experimental result (NDV: Avian pneumo-encephalitis virus, NC: negative control).
Fig. 2 real-time fluorescence detection method NDV HN chimeric gene primer specificity experimental result.
(a) real-time fluorescence amplification curve, (b) real-time fluorescence solubility curve
Fig. 3 fluorescent signal reaches the proliferation time corresponding to maximum value and the relation curve between Avian pneumo-encephalitis virus template ribonucleic acid extension rate logarithmic value.
(a) real-time fluorescence amplification curve, (b) function relation curve
Specific implementation method
Below in conjunction with accompanying drawing, be described in further detail specific embodiment of the invention method, advantage and disadvantage of the present invention will be more clear along with description.But these embodiments are only exemplary; any restriction is not formed to scope of the present invention; what those skilled in the art should understand that is; can modify to the details of technical solution of the present invention and form lower without departing from the spirit and scope of the present invention or replace, but these amendments and replacement all fall within the scope of protection of the present invention.
Embodiment
1. test materials and method
1.1 test materials
Inoculation Avian pneumo-encephalitis virus LaSota strain (preservation of equipment technology institute of China Inst. of Quarantine Inspection Sciences) is to SPF level chicken embryo, and 37 DEG C, humidity 55-75%, cultivate 48 hours.The allantoic fluid reclaimed, after measured after Hemagglutination titer, uses RNeasy Mini kit (QIAGEN), extracts virus total RNA.
1.2 design of primers
DNAMAN software is used to compare to NDV HN chimeric gene, select HN gene conserved regions fragment as target sequence, use online software PrimerExplorer V4 (http://primerexplorer.jp/e/) to design the loop-mediated isothermal amplification (LAMP) primer group of HN gene.HN gene loop-mediated isothermal amplification (LAMP) primer group, be made up of a pair outer primer (SEQ No.1, SEQ No.2), a pair inner primer (SEQ No.3, SEQ No.4) and a pair ring primer (SEQNo.5, SEQNo.6), its primer sequence is:
1.3 loop-mediated isothermal amplification
The cumulative volume of loop-mediated isothermal amplification system is 25 μ L, comprises HN primer mixed solution 6 μ L, and reaction premixed liquid 14 μ L, response sample 5 μ L, with DEPC water surrogate response sample in the test of negative control.After adding sample, reaction tubes is placed in ring mediated isothermal amplification fluorescence detecting system, arranging amplification program is: 1. 65 DEG C, 60min, 2. 98 DEG C-80 DEG C, 0.05 DEG C/s, 10min, fluorescent signal in system Real-Time Monitoring reaction tubes, by obtain isothermal duplication curve and solubility curve judge test-results.
The specific test of 1.4 primer mixed solutions
Respectively with ND LaSota vaccine strain (NDV vaccine strain LaSota) nucleic acid (preservation of equipment technology institute of China Inst. of Quarantine Inspection Sciences) and containing A/ Hangzhou/1/2013 (H7N9), A/ environment/Jiangxi/21/2011 (H9N2), Putuo/1203/2013, A/ Shanghai (H1N1 pdm09), Changning/1507/2012, A/ Shanghai (H3N2), in Haidian/1386/2013, B/ Beijing (Victoria) and B/ Chongqing/1361/2013 (Yamagata) strains of influenza viruses nucleic acid (being presented by China Sickness Prevention Control Center Virus Disease Prevention Control Institute with top plate) be template, real-time fluorescence systems axiol-ogy is carried out with HN gene loop-mediated isothermal amplification (LAMP) primer mixed solution, replace template as negative control using DEPC water simultaneously, the specificity of inspection primer mixed solution.
1.5 fluorescent signals reach the quantitative relationship analysis between proliferation time corresponding to maximum value and template ribonucleic acid extension rate logarithmic value
The newcastle disease virus RNA of extraction is carried out successively 10 times of gradient dilutions (1: 10,1: 100,1: 1000,1: 10000,1: 100000), by real-time fluorescence detection system, each doubling dilution RNA is detected respectively.The detected result of each reaction is carried out fluorescent signal in the logarithmic value of viral template RNA extension rate and real-time fluorescence detection method and is reached the linear analysis between the proliferation time of maximum value.
2. test-results
2.1 Avian pneumo-encephalitis virus chick embryo allantoic liquid Hemagglutination titer experimental results
The SPF chick embryo allantoic liquid not inoculating and inoculate Avian pneumo-encephalitis virus LaSota strain is carried out 1: 2 doubling dilution respectively, adopts 1% chicken red blood cell to carry out Hemagglutination titer experiment.As shown in Fig. 1 (NDV: Avian pneumo-encephalitis virus, NC: negative control), compared with control group, Avian pneumo-encephalitis virus is in dilution 2
7doubly chicken red blood cell can be made to condense completely, the Hemagglutination titer of this Avian pneumo-encephalitis virus is 128.
2.2 real-time fluorescence detection method primer specificity experimental results
Respectively with ND LaSota vaccine strain nucleic acid and containing A/ Hangzhou/1/2013 (H7N9), A/ environment/Jiangxi/21/2011 (H9N2), Putuo/1203/2013, A/ Shanghai (H1N1 pdm09), Changning/1507/2012, A/ Shanghai (H3N2), in Haidian/1386/2013, B/ Beijing (Victoria) and B/ Chongqing/1361/2013 (Yamagata) strains of influenza viruses nucleic acid is template, real-time fluorescence systems axiol-ogy is carried out with HN gene loop-mediated isothermal amplification (LAMP) primer mixed solution, test-results is as shown in (a) and (b) of Fig. 2, only have that ND LaSota vaccine strain is nucleic acid-templated occurs amplified signal, and solubility curve is single sharp peak, illustrate that the specificity of the loop-mediated isothermal amplification (LAMP) primer mixed solution of this HN gene is good.
2.3 fluorescent signals reach the quantitative analysis result between proliferation time corresponding to maximum value and template ribonucleic acid extension rate logarithmic value
The newcastle disease virus RNA of extraction is carried out successively 10 times of gradient dilutions (1: 10,1: 100,1: 1000,1: 10000,1: 100000), detected respectively each doubling dilution RNA by real-time fluorescence detection system, result is as shown in Fig. 3 (a).The quantitative relationship analytical results that Fig. 3 (b) (X-coordinate shows the logarithmic value of each reaction inner formword RNA extension rate, and ordinate zou is the proliferation time that each reaction fluorescent signal reaches maximum value) is Fig. 3 (a).Diluting HA gene ring mediated isothermal amplification linear equation in 1: 10-1: 100000 scopes at Avian pneumo-encephalitis virus masterplate RNA is y=1.3000x+6.1667, R
2the proliferation time that=0.9985, prompting HN gene by fluorescence signal reaches maximum value extends with the extension rate increase of reaction system RNA template, and linear relationship is good.1: 100000 template ribonucleic acid can be reached by the detection limit of the known the method HN gene of Fig. 3 (a).
Claims (6)
1. a set of loop-mediated isothermal amplification (LAMP) primer group detected for Avian pneumo-encephalitis virus, by hemagglutinin-neuraminidase (hemagglutinin-neuraminidase, HN) gene loop-mediated isothermal amplification (LAMP) primer group is formed, it is characterized in that, can be used for the loop-mediated isothermal amplification detecting Avian pneumo-encephalitis virus;
Described NDV HN chimeric gene loop-mediated isothermal amplification (LAMP) primer group, it is characterized in that, be made up of a pair outer primer (SEQ No.1, SEQ No.2), a pair inner primer (SEQ No.3, SEQ No.4) and a pair ring primer (SEQ No.5, SEQ No.6), its primer sequence is:
2. NDV HN chimeric gene loop-mediated isothermal amplification (LAMP) primer group as claimed in claim 1, it is characterized in that, the method detected for Avian pneumo-encephalitis virus comprises: 1) real-time fluorescence detection method, it is characterized in that, ring mediated isothermal amplification fluorescence detecting system is used to realize synchronous isothermal duplication and fluorescent signal in real time in detection reaction pipe, obtain isothermal duplication curve and solubility curve, and then judge detected result; 2) real-time turbidimetric assay, turbid ity signal in Real-Time Monitoring reaction tubes during reaction, obtains isothermal duplication curve, and then judges detected result; 3) uv irradiating fluorescence detection, irradiates under ultraviolet light by reaction solution in reaction tubes after amplified reaction, the test positive result of the fluorescence that takes on a red color, and what do not take on a red color is detected as negative findings; 4) direct fluorescence appearance method, it is characterized in that, in amplified reaction forward reaction pipe, add fluorexon as fluorescent indicator, by the direct result of determination of naked eyes, in reaction tubes, reaction solution is greeny is positive findings, and not greeny is negative findings; 5) gel electrophoresis assays, is characterized in that, having feature trapezoid-shaped strips person after reaction during reaction solution electrophoresis for positive findings, is negative findings without feature trapezoid-shaped strips person; 6) micro-total analysis method or micro-fluidic chip method or chip lab method, it is characterized in that, sample dissociation, nucleic acid extraction purifying, isothermal duplication and detection are all integrated on micro-total analysis system and automatically complete, result decision method comprises aforesaid real-time fluorescence detection method, in real time turbidimetric assay, uv irradiating fluorescence detection, directly fluorescence appearance method, gel electrophoresis assays, this micro-total analysis system is driven by micro diaphragm pump (valve), by unlatching and the closed drived control realizing microfluid sample of time variable control micro diaphragm pump (valve).
3. the test kit detected for Avian pneumo-encephalitis virus, it is characterized in that, form primarily of NDV HN chimeric gene loop-mediated isothermal amplification (LAMP) primer group solution as claimed in claim 1 and loop-mediated isothermal amplification premixed liquid, Avian pneumo-encephalitis virus positive quality control product and negative quality control product, adopt real-time fluorescence detection method to carry out the rapid detection of Avian pneumo-encephalitis virus;
Described NDV HN chimeric gene loop-mediated isothermal amplification (LAMP) primer group solution, it is characterized in that, configured according to a certain percentage by a pair inner primer, a pair outer primer and a pair ring primer to form, wherein inner primer and outer primer by 6: 1-12: 1 proportions become, preferred proportion is 8: 1, and the ratio of ring primer and inner primer is 1: 2;
Described loop-mediated isothermal amplification premixed liquid, is characterized in that, by Bst polysaccharase, dNTP, trimethyl-glycine, MgSO
4other reversed transcriptive enzymes as AMV reverse transcriptase or equivalence are formed with buffered soln and reversed transcriptive enzyme, comprise and amplified production can be made to send a kind of commercial kit Isothermal Master Mix of fluorescent signal or other test kits of equivalence;
Described Avian pneumo-encephalitis virus positive quality control product, is characterized in that, can be the total serum IgE extracted by the recombinant plasmid of Avian pneumo-encephalitis virus synthetic fragment structure, in-vitro transcription RNA or Avian pneumo-encephalitis virus LaSota strain.Described negative quality control product, is characterized in that, is the medium not containing newcastle disease virus gene pack section.
4. the using method of Avian pneumo-encephalitis virus detection kit as claimed in claim 3, it is characterized in that, respective rings mediated isothermality amplification primer sets solution 6 μ L is added respectively in HN gene loop-mediated isothermal amplification pipe, loop-mediated isothermal amplification premixed liquid 14 μ L, sample to be tested, positive quality control product or negative quality control product respectively add 5 μ L, ring mediated isothermal amplification fluorescence detecting system is used to realize synchronous isothermal duplication and fluorescent signal in real time in detection reaction pipe, obtain isothermal duplication curve and solubility curve, the amplification program of described fluorescence detecting system is: 1. 65 DEG C, 60min, 2. 98 DEG C-80 DEG C, 0.05 DEG C/s, 10min.
5. the using method of Avian pneumo-encephalitis virus detection kit as claimed in claim 3, it is characterized in that, analysis and the decision method of amplification comprise:
1) when there is platform in the amplification curve that ring mediated isothermal amplification fluorescence detecting system records, its fluorescent value is higher than background noise signal 5 times and the judgement that solubility curve is single sharp peak is positive findings, and its fluorescent value is higher than background noise signal 5 times but the judgement that solubility curve is not single sharp peak is nonspecific cross interference signal; Its fluorescent value is negative findings less than or equal to the judgement of background noise signal 5 times;
2) when sample to be tested HN gene loop-mediated isothermal amplification pipe is all negative findings, then Avian pneumo-encephalitis virus is not detected; Sample to be tested is positive findings, then have newcastle disease virus infection; The amplification curve of HN gene with reading during 15min for benchmark.
6. the using method of Avian pneumo-encephalitis virus detection kit as claimed in claim 3, it is characterized in that, during real-time fluorescence detects, each reaction fluorescent signal reaches between proliferation time corresponding to maximum value and the logarithmic value of template ribonucleic acid thinning ratio, be there is linear relationship, R in 1: 10-1: 100000 scopes in thinning ratio
2> 0.995, method detection limit can reach 1: 100000 Avian pneumo-encephalitis virus template ribonucleic acid.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106367537A (en) * | 2016-12-02 | 2017-02-01 | 刘文俊 | RT-LAMP visual kit for detecting goose newcastle disease virus (NDV) and application thereof |
| CN106979940A (en) * | 2017-04-17 | 2017-07-25 | 浙江大学 | A kind of device and method for carrying out nucleic acid amplification detection |
| CN110578019A (en) * | 2019-10-30 | 2019-12-17 | 广西壮族自治区兽医研究所 | Detection kit for distinguishing newcastle disease virulent strains and attenuated strains by double fluorescence LAMP (loop-mediated isothermal amplification) and primer group thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2007091580A1 (en) * | 2006-02-07 | 2007-08-16 | Nippon Biologicals, Inc. | Novel vaccine carrier |
| CN104099430A (en) * | 2014-08-01 | 2014-10-15 | 中国检验检疫科学研究院 | Ring mediate isothermal amplification kit for diagnosing H7N9 hypotype avian influenza virus and application method thereof |
| CN104561380A (en) * | 2014-12-17 | 2015-04-29 | 广州谱泰生物技术有限公司 | Synchronous detection kit and method for avian influenza and newcastle disease virus |
-
2015
- 2015-07-07 CN CN201510391606.7A patent/CN104911279A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2007091580A1 (en) * | 2006-02-07 | 2007-08-16 | Nippon Biologicals, Inc. | Novel vaccine carrier |
| CN104099430A (en) * | 2014-08-01 | 2014-10-15 | 中国检验检疫科学研究院 | Ring mediate isothermal amplification kit for diagnosing H7N9 hypotype avian influenza virus and application method thereof |
| CN104561380A (en) * | 2014-12-17 | 2015-04-29 | 广州谱泰生物技术有限公司 | Synchronous detection kit and method for avian influenza and newcastle disease virus |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106367537A (en) * | 2016-12-02 | 2017-02-01 | 刘文俊 | RT-LAMP visual kit for detecting goose newcastle disease virus (NDV) and application thereof |
| CN106367537B (en) * | 2016-12-02 | 2019-08-06 | 仲恺农业工程学院 | It is a kind of detect goose newcastle disease virus RT-LAMP visualizing agent box and its application |
| CN106979940A (en) * | 2017-04-17 | 2017-07-25 | 浙江大学 | A kind of device and method for carrying out nucleic acid amplification detection |
| CN110578019A (en) * | 2019-10-30 | 2019-12-17 | 广西壮族自治区兽医研究所 | Detection kit for distinguishing newcastle disease virulent strains and attenuated strains by double fluorescence LAMP (loop-mediated isothermal amplification) and primer group thereof |
| CN110578019B (en) * | 2019-10-30 | 2023-04-28 | 广西壮族自治区兽医研究所 | Double-fluorescence LAMP (loop-mediated isothermal amplification) detection kit for distinguishing strong and weak viruses of newcastle disease and primer group thereof |
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Application publication date: 20150916 |