CN108531629A - A kind of PCR amplification primer of quick detection Friedlander's bacillus and its application - Google Patents
A kind of PCR amplification primer of quick detection Friedlander's bacillus and its application Download PDFInfo
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Abstract
The invention discloses a kind of PCR amplification primer of quick detection Friedlander's bacillus and its applications, belong to animal bacteria and molecular biology field.The PCR amplification primer is by with SEQ ID NO:The sense primer of nucleotide sequence shown in 1 and have SEQ ID NO:The downstream primer of nucleotide sequence shown in 2 forms；Above-mentioned primer is used to prepare the PCR amplification kit of detection Friedlander's bacillus.The PCR detection method that the kit provides has the specificity and sensibility of height, it is reproducible, it is with a high credibility, specific detection goes out Friedlander's bacillus, fast and accurately obtain testing result, cheap, easy to operate simultaneously, suitable base uses, and can be used as a kind of quick, accurate, simplicity detection instrument of the quick discriminating in Friedlander's bacillus laboratory and extensive epidemiological survey.
Description
Technical field
The present invention relates to animal bacteria and technical field of molecular biology, particularly relate to a kind of quick, easy, low
Cost detects PCR amplification primer and its application of Friedlander's bacillus.
Background technology
Friedlander's bacillus（Klebsiella pneumonia, Kpn）It is blue for the leather of enterobacteriaceae Klebsiella
Family name's negative bacterium.The organs such as enteron aisle, the respiratory tract of humans and animals can be encroached on, so as to cause pneumonia, hepatapostema, wound infection and lose
The symptoms such as mass formed by blood stasis are a kind of cause of diseases of Amphixenosis.Meanwhile after cattle infected Friedlander's bacillus, to more killing property Pasteur bar
The neurological susceptibility of the cause of diseases such as bacterium, Mannheimia haemolytica and Mycoplasma bovis increases.Currently, the pathogen has in the world
Distribution, it is popular same very extensive in the distribution of China, huge economic loss is caused, the important disease for threatening ox aquaculture is become
One of original.
So far, the means report of the Friedlander's bacillus detected both at home and abroad is less.Mainly there is ELISA method, thin
The identification of bacterium culture of isolated Gram's staining, biochemical test etc..Not only time-consuming for these technologies, and detection is insensitive, and PCR method
Cause of disease can be not only diagnosed to be with rapid sensitive, but also for preclinical pathogen infection, specificity is stronger.
Invention content
The purpose of the present invention is to solve problems of the prior art, a kind of low cost, quickly and accurately is provided
Detect PCR amplification primer and its application of Friedlander's bacillus.Technical solution used in purpose is to realize the present invention：Packet
Include sample collection, the extraction of pathogenic genes group DNA, the design and optimization of specific primer, PCR amplification, agarose gel electrophoresis
Detect pcr amplification product and result judgement.
A kind of PCR amplification primer of quick detection Friedlander's bacillus, the PCR amplification primer are according to pneumonia gram
The phoE genes of the primary Salmonella of thunder are designed, and the target fragment size of amplification is 571bp, and the sequence of primer is respectively：
Forward primer：5’- CCATGCACTACTTCAGCGAT -3’ SEQ ID NO:1
Reverse primer：5’- CAGGCTTCCGCTTTCGAAC -3’ SEQ ID NO:2
Primer concentration is diluted to 25 pmol/ μ L using preceding.
Preferably, by the PCR amplimers in the application for preparing detection Friedlander's bacillus kit.
The present invention also provides a kind of PCR amplification kits of quick detection Friedlander's bacillus, including with SEQ ID
NO:The sense primer of nucleotide sequence shown in 1 and have SEQ ID NO:The downstream primer of nucleotide sequence shown in 2.
Preferably, further include 10 × PCR Buffer, dNTPs, ES-Taq archaeal dna polymerase and RNase-Free water,
PCR amplification method is established using the PCR amplification kit of quick detection Friedlander's bacillus, reaction system is established with 25 μ
L is counted,
10×PCR Buffer 5 μL
dNTPs（10mM each ） 2 μL
ES-Taq archaeal dna polymerases（5U/μL） 1 μL
Forward primer（25 pmol/μL） 1 μL
Reverse primer（25 pmol/μL） 1 μL
3 μ L of DNA profiling
RNase-Free water supply 25 μ L.
Preferably, 10 × PCR Buffer include KCL 20-30 mM, MgSO4 3-10 mM、10-20 mM
Tris-HCL。
PCR response procedures are 95 DEG C of 5 min in the preferred PCR amplification kit；Then carry out 35 95 DEG C
35 s, 58 DEG C of 40 s, the cycle of 72 DEG C of 45 s；10 min of last 72 DEG C of extensions.
A kind of PCR amplification kit of above-described quick detection Friedlander's bacillus, the result judgement are
The method detected using agarose gel electrophoresis：Pcr amplification product is subjected to electrophoresis on 1.5% Ago-Gel, is observed
Whether purposeful band.Positive control and negative control are set up first, if having amplified the specificity of 571 bp from sample
Band then illustrates that there are Friedlander's bacillus for the sample；If sample does not amplify the specific band of 571 bp, say
The bright sample does not contain Friedlander's bacillus.
The present invention substantive distinguishing features and significant progress be：
1）High specificity
The PCR detection method specific detection of the Friedlander's bacillus of the present invention goes out Friedlander's bacillus, the moon detected
Property control cause of disease such as Pyrogenes, Mannheimia haemolytica, Pasteurella, Mycoplasma bovis, 3 type of bovine parainfluenza virus, Niu Chuanran
Property rhinotracheitis virus, Escherichia coli and water compare no positive result.
2）High sensitivity
The PCR detection method high sensitivity of the Friedlander's bacillus of the present invention, minimum detection are limited to 3.88 × 10-4 ng/μL。
3）It takes less, is of low cost
Compared with being detected by Antigen isolation and identification, the PCR detection method of Friedlander's bacillus of the invention, the time at
This, workload etc. there is apparent advantage, interpretation of result judgement is carried out from nucleic acid extraction to agarose gel electrophoresis, can be
It is completed in 5 hours.
4）Accuracy is high, stability is good
With 3.88 × 101 ng/μL、3.88×100 ng/μL、3.88×10-1 ng/μL、3.88×10-2 ng/μL、3.88×
10-3、3.88×10-4 Ng/ μ L and 3.88 × 10-5 The recombinant plasmid standard sample of ng/ μ L is carried out at the same time PCR, is repeated 3 times respectively
Detection.As a result it shows that the result of 3 amplifications is consistent, shows that the reaction system for the PCR detection method established is reproducible.
Description of the drawings
Fig. 1 is annealing temperature screening test result：Wherein M：DNA marker 100 bp ladder 、1：50℃、2：52
℃、
3：54℃、4：56℃、5：58℃、6：60℃、7：62℃、8：Water compares.
Fig. 2 specific detection results：Wherein M：DNA marker 100bp ladder、1：Friedlander's bacillus, 2：It is hidden
Secret bacillus, 3：Mannheimia haemolytica, 4：Pasteurella, 5：Mycoplasma bovis, 6：3 type of bovine parainfluenza virus, 7：Ox infectiousness nose
Bronchitis virus, 8：Escherichia coli, 9：Water compares
Fig. 3 is sensitivity Detection result：Wherein M：DNA marker 100bp ladder、1：3.88×101ng/μL、2：
3.88×100 ng/μL、3：3.88×10-1 ng/μL、4：3.88×10-2 ng/μL、5：3.88×10-3 ng/μL、6 3.88×
10-4 ng/μL、7：3.88×10-5 ng/μL；8：Water compares.
Fig. 4 is clinical sample detection electrophoretogram：Wherein M：DNA marker 100bp ladder, wherein, swimming lane 1：It is positive
Control；2：Negative control；Swimming lane 4,5,7,9,12,14,15,18 is positive findings；Swimming lane 3,6,8,10,11,13,16,17,
19,20 be negative findings.
Specific implementation mode
The present invention program is described in further detail with reference to embodiment, following the description is merely to explain this hair
It is bright, its content is not defined.Experimental method used in following embodiments is conventional side unless otherwise specified
Method, material, reagent etc. used in following embodiments, is commercially available unless otherwise specified.
Embodiment 1 establishes the PCR method of quickly detection Friedlander's bacillus
1, the preparation of material
Friedlander's bacillus, Pyrogenes, Mannheimia haemolytica, Pasteurella, Mycoplasma bovis, 3 type of bovine parainfluenza virus,
Infectious bovine rhinotrachetis virus and Escherichia coli are that veterinary institute separation identification in Guangxi preserves, and tissue sample faces from animal doctor
Bed.10 × PCR Buffer, dNTPs, ES-Taq archaeal dna polymerase, DNA/RNA extracts kits, bacterial genomes DNA extractions
Kit is century bio tech ltd purchased from health.
2, the design and synthesis of PCR primer
Tetraploid rice analysis is carried out according to the Friedlander's bacillus phoE gene orders in GenBank, selects conserved sequence
Area goes out specificity amplification primer as amplification region, using 7.0 primer-design softwares of Oligo and BLAST software program designs,
The sequence of wherein primer is respectively：
Forward primer：5’- CCATGCACTACTTCAGCGAT -3’ SEQ ID NO:1
Reverse primer：5’- CAGGCTTCCGCTTTCGAAC -3’ SEQ ID NO:2
The target gene fragment size of amplification is 571 bp,
Upstream and downstream primer is using being preceding diluted to 25 pmol/ μ L.
3, the extraction of template DNA
Sample treatment：
（1）Pure bacterial cultures：It takes in right amount as in sterile centrifugation tube, then abandons supernatant after centrifuging, take precipitation；
（2）Tissue sample：It is ground first, multigelation, supernatant is taken after centrifugation.
Reuse the extraction of bacterial genomes DNA extraction kit.
4, PCR reaction systems are established
The PCR method of the quick detection Friedlander's bacillus, reaction system are established in terms of 25 μ L
10×PCR Buffer 5 μL
dNTPs（10mM each ） 2 μL
ES-Taq archaeal dna polymerases（5U/μL） 1 μL
Forward primer（25 pmol/μL） 1 μL
Reverse primer（25 pmol/μL） 1 μL
3 μ L of DNA profiling
RNase-Free water supply 25 μ L.
5, PCR response procedures
The response procedures of the PCR method of the quick detection Friedlander's bacillus, first choice carry out optimum annealing temperature determination
Experiment, after determining annealing temperature, the PCR response procedures used are 95 DEG C of 5 min；Then 35 95 DEG C of 35 s of progress, 58
DEG C 40 s, the cycle of 72 DEG C of 45 s；10 min of last 72 DEG C of extensions.
6, result judgement
The result judgement is the method using agarose gel electrophoresis detection：Agar by pcr amplification product 1.5%
Electrophoresis is carried out on sugared gel, sees whether purposeful band.Positive control and negative control are set up first, if expanded from sample
Increase the specific band for 571 bp, then illustrates that there are Friedlander's bacillus for the sample；If sample does not amplify 571
The specific band of bp then illustrates that the sample does not contain Friedlander's bacillus.
7, specific detection
With genomic DNA/RNA of the experiment strain of extraction and control strain（It is cDNA that RNA virus, which first carries out reverse transcription,）Carry out PCR expansions
Increase, examines the specificity of PCR method.
8, sensitivity Detection
The target fragment of the PCR amplification of Friedlander's bacillus is connect with PMD-18T carriers, conversion Escherichia coli DH5α, use
Small amount plasmid extraction agent box extracts plasmid, is identified through PCR, recombinant plasmid is served Hai Ying fine horses Bioisystech Co., Ltd and surveyed
Sequence determines.Positive recombinant plasmid is purified, after measuring initial concentration, with the continuous 10 times of doubling dilutions of RNA-Free Water, is used
The reaction condition of optimization carries out PCR amplification, carries out sensitivity Detection.
9, repeatability detection
With 3.88 × 101 ng/μL、3.88×100 ng/μL、3.88×10-1 ng/μL、3.88×10-2 ng/μL、3.88×
10-3、3.88×10-4 Ng/ μ L and 3.88 × 10-5 The recombinant plasmid standard sample of ng/ μ L is carried out at the same time PCR, is repeated 3 times respectively
Detection, examines the Stability and veracity of detection method.
10, clinical sample detects
50 parts of tissue samples of clinical acquisitions are ground respectively, multigelation, genomic DNA is extracted using kit, using building
Vertical PCR method is expanded.
Embodiment 2 quickly detects the annealing temperature experiment of Friedlander's bacillus PCR method
Respectively to 50 DEG C of annealing temperature, 52 DEG C, 54 DEG C, 56 DEG C, 58 DEG C, 60 DEG C, 62 DEG C of progress PCR amplifications, determine most
Good annealing temperature.The results show that the inclusiveness of designed primer pair annealing temperature is big, annealing temperature be 50 DEG C, 52
DEG C, 54 DEG C, 56 DEG C, 58 DEG C, 60 DEG C, under 62 DEG C of response procedures, can amplify single purpose band well
（Fig. 1）.For this purpose, the response procedures that this PCR method uses are：95 ℃ 5 min；Then 35 95 DEG C of 35 s of progress, 58 DEG C
40 s, the cycle of 72 DEG C of 45 s；Last 72 DEG C of 10 min extends.
Embodiment 3 detects the specific detection result of Friedlander's bacillus PCR method
Extract Friedlander's bacillus, Pyrogenes, Mannheimia haemolytica, Pasteurella, Mycoplasma bovis, bovine parainfluenza virus
3 types, infectious bovine rhinotrachetis virus and genomic DNA/RNA of Escherichia coli（It is cDNA that RNA, which carries out reverse transcription,）, use is excellent
The reaction system and response procedures changed carry out PCR amplification, detect the specificity of detection method, the results show that only
Friedlander's bacillus sample amplification has gone out target fragment band, is positive findings, 7 plants of control strain reaction tubes and water control reaction
Amplified band is not detected in Guan Jun, is negative findings（Fig. 2）, show that this method has specificity well.
Embodiment 4 detects the sensitivity Detection result of Friedlander's bacillus PCR method
The target fragment of the PCR amplification of Friedlander's bacillus is connect with PMD-18T carriers, conversion Escherichia coli DH5α, use
Small amount plasmid extraction agent box extracts plasmid, is identified through PCR, recombinant plasmid is served Hai Ying fine horses Bioisystech Co., Ltd and surveyed
Sequence determines.Positive recombinant plasmid is purified, it is 3.88 × 10 to measure initial concentration1It is continuous with RNA-Free Water after ng/ μ L
10 times of doubling dilutions carry out PCR amplification using the reaction condition of optimization, carry out sensitivity Detection, the results show that the PCR established
Detection method lowest detection is limited to 3.88 × 10-4 ng/μL（Fig. 3）.
The Stability and veracity testing result of 5 Friedlander's bacillus PCR method of embodiment
With 3.88 × 101 ng/μL、3.88×100 ng/μL、3.88×10-1 ng/μL、3.88×10-2 ng/μL、3.88×
10-3、3.88×10-4 Ng/ μ L and 3.88 × 10-5 The standard sample of ng/ μ L is carried out at the same time PCR amplification, is repeated 3 times inspection respectively
It surveys.The results show that reproducible results is good, show that the PCR detection method established is reproducible, stability is high.
The Preliminary Applications of 6 Friedlander's bacillus PCR method of embodiment
50 parts of tissue samples of clinical acquisitions are ground respectively, multigelation, extracts genomic DNA using kit, respectively PCR
Amplification.Fig. 4 shows the testing result of wherein 18 parts samples, the results showed that, there are 8 parts of sample detections to go out the spy of Friedlander's bacillus
Anisotropic band is positive findings.
The above description is merely a specific embodiment, but scope of protection of the present invention is not limited thereto, any
The change or replacement expected without creative work, should be covered by the protection scope of the present invention.Therefore, of the invention
Protection domain should be determined by the scope of protection defined in the claims.
Sequence table
<110>Veterinary Institute of Guangxi Zhuang Autonomous Region
<120>A kind of PCR amplification primer of quick detection Friedlander's bacillus and its application
<141> 2018-05-30
<160> 2
<170> SIPOSequenceListing 1.0
<210> 1
<211> 20
<212> DNA
<213>Artificial sequence (Artificial sequence Latin)
<400> 1
ccatgcacta cttcagcgat 20
<210> 2
<211> 19
<212> DNA
<213>Artificial sequence (Artificial sequence Latin)
<400> 2
caggcttccg ctttcgaac 19
Claims (8)
1. a kind of PCR amplification primer of quick detection Friedlander's bacillus, which is characterized in that the PCR amplification primer is by having
There are SEQ ID NO:The sense primer of nucleotide sequence shown in 1 and have SEQ ID NO:The downstream of nucleotide sequence shown in 2
Primer forms.
2. quickly detecting the PCR amplification primer of Friedlander's bacillus according to claim 1, which is characterized in that described
PCR amplification primer is that the phoE genes of foundation Friedlander's bacillus are designed, and the target fragment size of amplification is 571 bp.
3. quickly detecting the PCR amplification primer of Friedlander's bacillus according to claim 1, which is characterized in that described
PCR amplification primer is in the application for preparing detection Friedlander's bacillus kit.
4. a kind of PCR amplification kit quickly detecting Friedlander's bacillus as described in claim 1, which is characterized in that packet
It includes with SEQ ID NO:The sense primer of nucleotide sequence shown in 1 and have SEQ ID NO:Nucleotide sequence shown in 2
Downstream primer.
5. quickly detecting the PCR amplification kit of Friedlander's bacillus according to claim 4, which is characterized in that also wrap
Include 10 × PCR Buffer, dNTPs, ES-Taq archaeal dna polymerase and RNase-Free water.
6. quickly detecting the PCR amplification kit of Friedlander's bacillus according to claim 5, which is characterized in that described
10 × PCR Buffer include KCL 20-30 mM, MgSO4 3-10 mM、10-20 mM Tris-HCL。
7. the PCR amplification kit of the quick detection Friedlander's bacillus according to claim 4 or 5, which is characterized in that institute
State the SEQ ID NO of PCR amplification kit:1 primer and SEQ ID NO:2 primers use a concentration of 25 pmol/ μ L.
8. the PCR amplification kit of the quick detection Friedlander's bacillus according to claim 4 or 5, which is characterized in that institute
PCR response procedures are 95 DEG C of 5 min in the PCR amplification kit stated；Then 35 95 DEG C of 35 s of progress, 58 DEG C 40
S, the cycle of 72 DEG C of 45 s；10 min of last 72 DEG C of extensions.
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109628621A (en) * | 2019-02-02 | 2019-04-16 | 广西壮族自治区兽医研究所 | A kind of real-time quantitative LAMP primer group and kit detecting Friedlander's bacillus |
| CN110499374A (en) * | 2019-08-12 | 2019-11-26 | 山东农业大学 | A kind of triple PCR primer sets, kit and application detecting Klebsiella Pneumoniae |
| CN110669852A (en) * | 2019-11-04 | 2020-01-10 | 深圳市人民医院 | Kit for detecting high-toxicity non-mucus Klebsiella pneumoniae |
| CN111206110A (en) * | 2020-03-25 | 2020-05-29 | 台州学院 | Specific primers, kit and method for synchronously detecting klebsiella pneumoniae and golden yellow bacillus of large yellow croaker source |
| CN111500751A (en) * | 2020-04-10 | 2020-08-07 | 刘洋 | Detection method and kit for rapidly detecting high-toxicity Klebsiella pneumoniae |
| CN112725489A (en) * | 2021-03-05 | 2021-04-30 | 山西大学 | Klebsiella pneumoniae characteristic sequence and qPCR rapid detection system and method thereof |
| CN114836550A (en) * | 2021-11-10 | 2022-08-02 | 江汉大学 | MNP (MNP) marker site of klebsiella pneumoniae, primer composition, kit and application of MNP marker site and primer composition |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2002086153A1 (en) * | 2001-04-24 | 2002-10-31 | Diatech Pty Ltd | DETERMINING RESISTANCE TO A $G(b)-LACTAM-CONTAINING ANTIBIOTIC BY COMPARING THE RATIO OF TWO $G(b)-LACTAMASE GENES |
| CN104531887A (en) * | 2015-01-15 | 2015-04-22 | 山东省动物疫病预防与控制中心 | Identification method for Klebsiella pneumoniae and adopted primers |
| CN104946762A (en) * | 2015-06-17 | 2015-09-30 | 中生北控生物科技股份有限公司 | Kit for detecting klebsiella pneumoniae |
-
2018
- 2018-05-31 CN CN201810543504.6A patent/CN108531629A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2002086153A1 (en) * | 2001-04-24 | 2002-10-31 | Diatech Pty Ltd | DETERMINING RESISTANCE TO A $G(b)-LACTAM-CONTAINING ANTIBIOTIC BY COMPARING THE RATIO OF TWO $G(b)-LACTAMASE GENES |
| CN104531887A (en) * | 2015-01-15 | 2015-04-22 | 山东省动物疫病预防与控制中心 | Identification method for Klebsiella pneumoniae and adopted primers |
| CN104946762A (en) * | 2015-06-17 | 2015-09-30 | 中生北控生物科技股份有限公司 | Kit for detecting klebsiella pneumoniae |
Non-Patent Citations (3)
| Title |
|---|
| 刘俊,杨忠，王翀主编: "《可食性包装材料质量检验》", 30 April 2017, 中国质检出版社 * |
| 张诗渝: ""肺炎克雷伯杆菌的分离鉴定及其对小鼠的致病性研究"", 《中国优秀硕士学位论文全文数据库 农业科技辑》 * |
| 李燕，张健: "《生物学实用实验技术分子》", 31 December 2011, 西安：第四军医大学出版社 * |
Cited By (13)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109628621A (en) * | 2019-02-02 | 2019-04-16 | 广西壮族自治区兽医研究所 | A kind of real-time quantitative LAMP primer group and kit detecting Friedlander's bacillus |
| CN109628621B (en) * | 2019-02-02 | 2022-02-15 | 广西壮族自治区兽医研究所 | Real-time quantitative LAMP primer group and kit for detecting Klebsiella pneumoniae |
| CN110499374A (en) * | 2019-08-12 | 2019-11-26 | 山东农业大学 | A kind of triple PCR primer sets, kit and application detecting Klebsiella Pneumoniae |
| CN110499374B (en) * | 2019-08-12 | 2020-07-17 | 山东农业大学 | Triple PCR primer group for detecting Klebsiella pneumoniae, kit and application |
| CN110669852A (en) * | 2019-11-04 | 2020-01-10 | 深圳市人民医院 | Kit for detecting high-toxicity non-mucus Klebsiella pneumoniae |
| CN111206110B (en) * | 2020-03-25 | 2023-04-07 | 台州学院 | Specific primers, kit and method for synchronously detecting klebsiella pneumoniae and golden yellow bacillus of large yellow croaker source |
| CN111206110A (en) * | 2020-03-25 | 2020-05-29 | 台州学院 | Specific primers, kit and method for synchronously detecting klebsiella pneumoniae and golden yellow bacillus of large yellow croaker source |
| CN111500751A (en) * | 2020-04-10 | 2020-08-07 | 刘洋 | Detection method and kit for rapidly detecting high-toxicity Klebsiella pneumoniae |
| CN111500751B (en) * | 2020-04-10 | 2023-10-27 | 刘洋 | Detection method and kit for rapidly detecting high-virulence klebsiella pneumoniae |
| CN112725489A (en) * | 2021-03-05 | 2021-04-30 | 山西大学 | Klebsiella pneumoniae characteristic sequence and qPCR rapid detection system and method thereof |
| CN112725489B (en) * | 2021-03-05 | 2022-09-23 | 山西大学 | Klebsiella pneumoniae characteristic sequence and qPCR rapid detection system and method thereof |
| CN114836550B (en) * | 2021-11-10 | 2023-06-16 | 江汉大学 | MNP (MNP) marking site of klebsiella pneumoniae, primer composition, kit and application of MNP marking site |
| CN114836550A (en) * | 2021-11-10 | 2022-08-02 | 江汉大学 | MNP (MNP) marker site of klebsiella pneumoniae, primer composition, kit and application of MNP marker site and primer composition |
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Application publication date: 20180914 |