CN106636428A - PCR (Polymerase Chain Reaction) method for quickly detecting and identifying serratia - Google Patents
PCR (Polymerase Chain Reaction) method for quickly detecting and identifying serratia Download PDFInfo
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Abstract
The invention provides a PCR (Polymerase Chain Reaction) method for quickly detecting and identifying serratia. In the method, a forward primer F: 5'-GATCCACCCCAACGTGTTCT-3' and a downstream primer R: 5'-GGCACTTTCACCGAAACCAC-3' are adopted to carry out PCR reaction. The PCR reaction is circularly set as follows: at 95DEG C, predegeneration is carried out for 2 min; circulation is carried out for 30 times at the temperature of 95DEG C for 30s, at the temperature of 56DEG C for 30s and at the temperature of 72DEG C for 30s; finally, extension is carried out for 5min at the temperature of 72DEG C, and reaction is finished. The method has the characteristics of high speed, high sensitivity, high specificity and the like for detecting and identifying the serratia, provides a technical means for detecting and identifying the serratia marcescens and the serratia liquefaciens of three classes of pathogenic microorganisms, simultaneously provides a method for quickly detecting and diagnosing the serratia and other enterobacteriaceae bacteria, and has an important meaning for the early diagnosis and identification of infection caused by the serratia.
Description
Technical field
Present invention design molecular Biological Detection identification field, and in particular to a kind of quick detection identification Serratia
PCR method.
Background technology
Husky thunder bacterium belongs to enterobacteriaceae in classification( Enterobacteriaceae), Serratia（Serratia）.Sha Lei
Pseudomonas is existing including Serratia marcescans（S.marcescens）, Serratia fonticola（S.fonticola）, Serratia liquefaciens（S. liquefaciens）Deng 14 kinds, two subspecies.In medical domain, each kind of Serratia mainly causes Hospital to really feel
Dye.Wherein Serratia marcescans and Serratia liquefaciens are put into the formulation of the Ministry of Public Health of China《The pathogenic microorganism register that the human world is infected》
(2006), the extent of injury belongs to the 3rd class.Husky thunder bacterium is distributed widely in enteron aisle and the breathing of water, soil, plant, the mankind and animal
It is the conditioned pathogen in air in road.The bacterium is considered as a long time harmless, but due to its have it is aggressive and right
Many commonly-used antibiosis have drug resistance, it has also become a kind of important pathogen, can cause pneumonia, urethral infection, the sepsis of people
Disease and Postoperative infection etc..In veterinary applications, husky thunder bacterium is the pathogen for causing garget and many animals disease, can be drawn
Play ox（Mammitis）And horse（Conjunctivitis）, goat（Sepsis）, chicken, many animals infection such as pig and rabbit, animal can pass through quilt
The various kinds of media such as the drinking-water of pollution, bedding and padding, apparatus and the spittle, dust, excrement comes in contact infection.In addition, in recent years drinking-water and
The pollution of husky thunder bacterium in dairy products is also increasingly taken seriously with existence.
We have found during the routine testing that import animalsderived feedstuffs are carried out with detection of Salmonella, are carrying out selecting mild-natured
When plate is separately cultured detection of Salmonella, Serratia fonticola is mainly to disturb one of bacterium.Carried out according to the biochemical characteristic of husky thunder bacterium a series of
Biochemistry and serological Identification conventional method it is time-consuming and workload is big, if can be after selective plate isolation, to doubting
The rapid differential diagnosis of enteron aisle detection of Salmonella and Serratia are carried out using molecular biology method like detection of Salmonella bacterium colony, undoubtedly will be carried
Detection of Salmonella detection operating efficiency in the inward feedstuff of height, greatly shortens subsequent biochemical and the time needed for serological Identification, section
Human-saving and material resources.At present, the molecular Biological Detection technical research for detection of Salmonella is relatively broad, there is various commercial reagents
Box is alternative, but there is no more perfect molecular Biological Detection means for Serratia.A kind of PCR that the present invention is provided is fast
The method of fast Testing and appraisal Serratia, it will help meet clinical medicine detection need of work and port goods is tested and puts quick inspection
Need of work is surveyed, with most important theories meaning and actual application value.
The content of the invention
It is an object of the invention to provide a kind of method that PCR quick detections identify Serratia, the present invention is for husky thunder
Bacterium tradition isolation and identification method cannot meet clinical medicine detection need of work and cannot meet port goods and test puts quick detection
Need of work these deficiencies conduct a research, it is desirable to provide a kind of quick, sensitivity is high, the detection method of high specificity is to husky thunder bacterium
Carry out Testing and appraisal.
The invention discloses a kind of method that PCR quick detections identify Serratia, sub- single with B in coding DNA gyrase
The gyrB genes of position albumen are target gene, and the molecular biological characteristics for designing primer pair Serratia are identified；According to
GenBank announce gene order design primer be：
Upstream primer is F：5 '-GATCCACCCCAACGTGTTCT-3 ',
Downstream primer is R：5’-GGCACTTTCACCGAAACCAC-3’.
The PCR reaction systems cumulative volume of the present invention is 25 μ L, wherein 2 × GoTaq Green Master Mix（The U.S.
Promega companies produce）12.5 μ L, each 1 μ L of upstream and downstream primer（Final concentration of 0.8 μM）, sterilize ddH2The μ L of O 9.5, DNA moulds
The μ L of plate 1.
The PCR reaction cycles of the present invention are set to：95 DEG C, the min of denaturation 2；95 DEG C of 30s, 56 DEG C of 30s, 72 DEG C
30s, 30 circulations；5 min of last 72 DEG C of extensions, reaction terminates.
It is of the invention to have the advantage that compared with the method for traditional separation identification：The inventive method is being examined to husky thunder bacterium
The features such as there is high quick, sensitivity, high specificity is surveyed in identification.The present invention is formulated for the Ministry of Public Health of China《What the human world was infected
Pathogenic microorganism register》(2006) detection of three class pathogenic microorganism Serratia marcescans and Serratia liquefaciens and identification in provides high
The technological means of effect, while contributing to realizing the rapid differential diagnosis of Serratia and other enterobacteriaceae lactobacteriaceaes, solves similar
The various practical problems of the rapid differential diagnosis to enteron aisle detection of Salmonella and Serratia are realized described in background technology, with potential
Actual application value.
A kind of quick detection provided by the present invention identifies the PCR method of Serratia, has the advantages that：
1st, high specificity：The specificity that the gyrB genes of B subunit proteins are designed as target gene with coding DNA gyrase is drawn
Thing, in PCR tests only it is amplifiable go out husky thunder bacterium, and including Salmonella typhimurtum, Salmonella enteritidis, Escherichia coli, Freund lemon
Lemon acidfast bacilli, Song Shi shigella dysenteriaes, Klebsiella oxytoca, Klebsiella Pneumoniae, enterococcus faecalis, clostridium perfringen etc. are at interior 9 kinds
The amplification of Fei Shalei bacterium is feminine gender, it is shown that the good specificity of the primer.
2nd, sensitivity is high：Enter performing PCR with the genomic DNA of husky thunder bacterium and react its detection sensitivity to can reach 9.785pg/ μ
L。
3rd, reliable results：Serratia rubidaea, Serratia ficaria, Serratia fonticola, serratia plymouthensis, glutinous is used
The 7 planting sand thunder bacterium such as matter sand thunder bacterium, Serratia liquefaciens, Serratia proteamaculans are verified to primer, therefore can fully ensure that knot
The reliability of fruit.
4th, practicality is good：The detection time according to needed for the biochemical characteristic and serological traditional isolation and identification method of bacterium
Long, the present invention compensate for the deficiency of traditional isolation and identification method using the method for molecular biology, greatly reduce detection work
When the timely diagnosis of the urgent patients such as required time, the septicemia caused by clinical Yin Shalei bacterium and treatment provide valuable
Between.Meanwhile, it is that the quick discriminating of enteron aisle detection of Salmonella and Serratia is examined in terms of the detection of Salmonella quarantine of import animalsderived feedstuffs
It is disconnected to provide new method.
Description of the drawings
Fig. 1 is Serratia PCR primer specific test result.Wherein, Fig. 1（a）Middle M is 100bp DNA ladder scale designations
Thing (be followed successively by from the bottom up 100bp, 200bp, 300bp, 400bp, 500bp, 600bp, 700bp, 800bp, 1000bp,
1500bp)；NC is negative control；Swimming lane 1～7 is followed successively by：Serratia fonticolaSerratia fonticola；Serratia proteamaculansSerratia proteamaculans；Serratia marcesensSerratia marcescens；Serratia ficariaSerratia ficaria；Serratia plymouthensisSerratia plymuthica；Serratia liquefaciensSerratia liquefaciens；
Serratia rubidaeaSerratia rubidaea；Fig. 1（b）Middle M (is followed successively by from the bottom up for 100bp DNA ladder marker things
100bp、200bp、300bp、400bp、500bp、600bp、700bp、800bp、1000bp、1500bp)；NC is negative control；
PC is positive control（Serratia fonticolaSerratia fonticola）；Swimming lane 1～9 is followed successively by：Escherichia coliEschericha coli；Citrobacter freundiiCitrobacter freundii；Song Shi shigella dysenteriaesShigella sonnei；Produce sour Cray primary
Bacterium Klebsiella oxytoca；Enterococcus faecalisEnterrococcus faecalis；Clostridium perfringen Enterobacter aerogenes；Salmonella enteritidis Salmonella enteritidis；Salmonella typhimurtum Salmonella typhimurium；Klebsiella Pneumoniae Klebsiella pneumoniae。
Fig. 2 is Serratia PCR primer optimum annealing temperature result of the test.Wherein：M is 100bp DNA ladder scale designation things
(be followed successively by from the bottom up 100bp, 200bp, 300bp, 400bp, 500bp, 600bp, 700bp, 800bp, 1000bp,
1500bp)；NC is negative control；The temperature of swimming lane 1～10 is followed successively by：53.2℃、53.9℃、55.0℃、56.2℃、 57.4
DEG C, 58.6 DEG C, 59.8 DEG C, 61.0 DEG C, 62.1 DEG C and 62.8 DEG C.
Fig. 3 is Serratia（Serratia fonticolaSerratia fonticola）PCR tests LDL result of the test.
Wherein：M be 100bp DNA ladder marker things (be followed successively by from the bottom up 100bp, 200bp, 300bp, 400bp, 500bp,
600bp、700bp、800bp、1000bp、1500bp)；NC is negative control；The temperature of swimming lane 1～8 is followed successively by：97.85 ng、
9.785 ng, 978.5 pg, 97.85 pg, 9.785 pg, 978.5 fg, 97.85 fg and 9.785 fg.
Specific embodiment
Embodiment 1：Serratia PCR primer specific test
The preparation of 1.1 materials
It is as follows for the husky thunder bacterium of examination：Serratia fonticolaSerratia fonticola;Serratia proteamaculansSerratia proteamaculans;Serratia marcesensSerratia marcescens;Serratia ficariaSerratia ficaria;
Serratia plymouthensisSerratia plymuthica;Serratia liquefaciensSerratia liquefaciens;Dark red husky thunder
BacteriumSerratia rubidaea；Fei Shalei bacterium are as follows：Escherichia coliEschericha coli;Citrobacter freundiiCitrobacter freundii;Song Shi shigella dysenteriaesShigella sonnei;Klebsiella oxytoca Klebsiella oxytoca;Enterococcus faecalisEnterrococcus faecalis;Clostridium perfringen Enterobacter aerogenes;Enteritis
Detection of Salmonella Salmonella enteritidis;Salmonella typhimurtum Salmonella typhimurium;Kerekou pneumonia primary
Bacterium Klebsiella pneumoniae。
Bacterial strain used above is all from domestic authority's Culture Collection.
The foundation of 1.2 PCR methods
1.2.1 the design synthesis of primer：GyrB based on husky thunder bacterium（The gene of B subunit proteins in coding DNA gyrase）Base
Because of sequence, by a large amount of the design for obtaining highly conserved and high specificity sequence for specific primer, Jing are compared and screen
The specificity of primer designed by NCBI Blast comparisons, by Shanghai, Sheng Gong biotechnologys Services Co., Ltd synthesizes.It is set
Meter primer sequence is as follows, and gropes its optimum annealing temperature for designed primer thermograde PCR instrument, sees embodiment 2.
Upstream primer F：5’ -GATCCACCCCAACGTGTTCT-3’
Downstream primer R：5’ -GGCACTTTCACCGAAACCAC-3’
Amplified fragments size is 279bp.
1.2.2 extracting genome DNA
1.2.2.1 boiling method：
1st, the streak inoculation of picking single bacterium colony is in LB agar plates, 37 DEG C of culture 18h-24h；
2nd, scrape 1～2 oese lawn to add in 150 microlitres of Tris-EDTA (TE) buffer solutions, mix, in 100 DEG C of boiling water baths
Boil 10min；
3rd, 12000 r/min centrifugations 10min, takes supernatant, and -20 DEG C save backup.
1.2.2.2 isolation kit method：Operate according to TIANGEN bacterial genomes DNA extraction kits operation instructions
Extract DNA.The DNA of bacteria for being extracted is stored in TE buffer solutions, with ultramicron nucleotides analyzer DNA concentration and matter are determined
Amount（OD260/OD280Between 1.7 ~ 1.9 and concentration be more than 20ng/ μ L when, can be used for follow-up PCR reaction）, -20 DEG C of preservations are standby
With.
1.2.3 pcr amplification reaction system：Cumulative volume be 25 μ L, the μ L of 2 × GoTaq Green Master Mix 12.5,
The each 1 μ L of upstream and downstream primer（Final concentration of 0.8 μM）, sterilize ddH2The μ L of O 9.5, the μ L of DNA profiling 1.
1.2.4 PCR reaction cycles are arranged：95 DEG C of min of denaturation 2；95 DEG C of 30s, 56 DEG C of 30s, 72 DEG C of 30s, 30
Individual circulation；72 DEG C of 5 min of extension, reaction terminates.
1.2.5 pcr amplification product detection and identification：6 μ LPCR products point samples are taken in 1.5% Ago-Gel（Second containing bromination
Ingot）, Ago-Gel is placed in 1 × TAE buffer solutions with after the min of 120V electrophoresis 50, take out, it is placed in gel imaging instrument
Middle uviol lamp is observed.If there is amplified band in the position of 279bp, expanded sample is illustrated for husky thunder bacterium.Electrophoresis is tied
Fruit shows that the pcr amplification product for there was only husky thunder bacterium has bright amplified band (see Fig. 1 in the position of 279bp（a）), it is non-with other
The DNA of husky thunder bacterium enters performing PCR reaction and does not observe there is amplified band (see Fig. 1 as template（b）), illustrate the specificity of primer
It is very strong.
Embodiment 2：PCR primer optimum annealing temperature is screened
2.1 materials prepare
The husky thunder bacterium extracted using 1.2.2 in embodiment 1（By taking Serratia fonticola as an example）The template that DNA reacts as PCR.
2.2 PCR
2.2.1 pcr amplification reaction system：Cumulative volume is 25 μ L, the μ L of 2 × GoTaq Green Master Mix 12.5, is implemented
The each 1 μ L of upstream and downstream primer described in 1.2.1 in example 1（Final concentration of 0.8 μM）, sterilize ddH2The μ L of O 9.5, the μ L of DNA profiling 1.
2.2.2 PCR reaction cycles are set to：95 DEG C of min of denaturation 2；95 DEG C of 30s, using grads PCR instrument 10 are arranged
Individual annealing temperature 53.2 DEG C～62.8 DEG C 30s, 72 DEG C of 30s, 30 circulations；72 DEG C of 5 min of extension, reaction terminates.
2.2.3 pcr amplification product detection and identification：6 μ LPCR products point samples are taken in 1.5% Ago-Gel（Second containing bromination
Ingot）, Ago-Gel is placed in 1 × TAE buffer solutions with after 120V electrophoresis 50min, take out, in being placed in gel imaging instrument
Observe under uviol lamp.There is amplified band on the position of 279bp（See Fig. 2）, wherein the amplified band brightness of swimming lane 1～5 without
Notable difference, the amplified band brightness of swimming lane 6～10 gradually dies down, while considering that properly increasing annealing temperature can improve specifically
Property amplification, so annealing temperature select be 56 DEG C.
Embodiment 3：Serratia PCR detection sensitivities are tested
3.1 materials prepare
The husky thunder bacterium for being extracted 1.2.2 in embodiment 1 using 10 times of concentration series dilution methods（By taking Serratia fonticola as an example）DNA
Stoste（ 97.85ng/μL）It is diluted to 9.785 ng, 978.5 pg, 97.85 pg, 9.785 pg, 978.5 fg, 97.85 fg
With 9.785 fg totally 7 variable concentrations gradients.Performing PCR sensitivity is entered to the husky thunder bacterium DNA together with stoste in interior totally 8 concentration
Test.
3.2 PCR are detected
3.2.1 pcr amplification reaction system：Cumulative volume is 25 μ L, the μ L of 2 × GoTaq Green Master Mix 12.5, is implemented
The each 1 μ L of upstream and downstream primer described in 1.2.1 in example 1（Final concentration of 0.8 μM）, sterilize ddH2The μ L of O 9.5, the μ L of DNA profiling 1.
3.2.2 PCR reaction cycles are arranged：95 DEG C of min of denaturation 2；95 DEG C of 30s, 56 DEG C of 30s, 72 DEG C of 30s, 30
Individual circulation；72 DEG C of 5 min of extension, reaction terminates.
3.2.3 pcr amplification product detection and identification：6 μ LPCR products point samples are taken in 1.5% Ago-Gel（Second containing bromination
Ingot）, Ago-Gel is placed in 1 × TAE buffer solutions with after 120V electrophoresis 50min, take out, in being placed in gel imaging instrument
Observe under uviol lamp.If there is amplified band in the position of 279bp, illustrating the sample of the concentration can be amplified.Electrophoresis
Still there is amplified band the position of 279bp when as a result showing that DNA concentration is 9.785 pg/ μ L（See Fig. 3）, illustrate this detection method
With higher sensitivity, 9.785 pg/ μ L can be reached.
The above is arranged for reaction ideal after present invention optimization, every according to scope of the present invention patent institute
The impartial change done and modification, should all belong to the covering scope of the present invention.
SEQUENCE LISTING
<110>Inspection ＆ Quarantine Technology Center of Fujian Entry-Exit Inspection ＆ Quar
<120>A kind of quick detection identifies the PCR method of Serratia
<130> 2
<160> 2
<170> PatentIn version 3.3
<210> 1
<211> 20
<212> DNA
<213>Artificial sequence
<400> 1
gatccacccc aacgtgttct 20
<210> 2
<211> 20
<212> DNA
<213>Artificial sequence
<400> 2
ggcactttca ccgaaaccac 20
Claims (3)
1. a kind of quick detection identifies the PCR primer of Serratia, it is characterised in that：The primer sequence is：Upstream primer is
F：5 '-GATCCACCCCAACGTGTTCT-3 ', downstream primer is R：5’-GGCACTTTCACCGAAACCAC-3’.
2. a kind of quick detection identifies the PCR method of Serratia, it is characterised in that：Entered using the primer described in claim 1
Performing PCR reacts, and PCR reaction systems cumulative volume is 25 μ L, wherein the μ L of 2 × GoTaq Green Master Mix 12.5, upstream and downstream
The each 1 μ L of primer and final concentration of 0.8 μM, sterilizing ddH2The μ L of O 9.5, the μ L of DNA profiling 1.
3. a kind of quick detection according to claim 2 identifies the PCR method of Serratia, it is characterised in that：PCR reacts
Be circularly set for：95 DEG C, the min of denaturation 2；95 DEG C of 30s, 56 DEG C of 30s, 72 DEG C of 30s, 30 circulations；Last 72 DEG C are prolonged
5 min are stretched, reaction terminates.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112029830A (en) * | 2020-09-16 | 2020-12-04 | 上海上药第一生化药业有限公司 | Method for improving sensitivity of PCR (polymerase chain reaction) for identifying pig, cattle and sheep derived components in protein biochemical raw material crude product |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101191146A (en) * | 2006-11-28 | 2008-06-04 | 佳能株式会社 | Primer for bacterium genome amplification reaction |
| CN101558168A (en) * | 2006-10-03 | 2009-10-14 | 国立大学法人岐阜大学 | Detection of bacterium by utilizing DnaJ gene and use thereof |
| CN101659992A (en) * | 2009-03-20 | 2010-03-03 | 曹际娟 | Detection kit and detection method for 5 new pathogens in foods |
| CN105018601A (en) * | 2015-06-26 | 2015-11-04 | 华侨大学 | Post-neurosurgical intracranial bacterial infection common pathogenic bacterium MLPA detection probe and application thereof |
| CN105368961A (en) * | 2015-12-15 | 2016-03-02 | 天津脉络生物科技有限公司 | Shewanella oneidensis quantitative detection fluorescent PCR reagent kit and application thereof |
-
2017
- 2017-01-23 CN CN201710048877.1A patent/CN106636428B/en active Active
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101558168A (en) * | 2006-10-03 | 2009-10-14 | 国立大学法人岐阜大学 | Detection of bacterium by utilizing DnaJ gene and use thereof |
| CN101191146A (en) * | 2006-11-28 | 2008-06-04 | 佳能株式会社 | Primer for bacterium genome amplification reaction |
| CN101659992A (en) * | 2009-03-20 | 2010-03-03 | 曹际娟 | Detection kit and detection method for 5 new pathogens in foods |
| CN105018601A (en) * | 2015-06-26 | 2015-11-04 | 华侨大学 | Post-neurosurgical intracranial bacterial infection common pathogenic bacterium MLPA detection probe and application thereof |
| CN105368961A (en) * | 2015-12-15 | 2016-03-02 | 天津脉络生物科技有限公司 | Shewanella oneidensis quantitative detection fluorescent PCR reagent kit and application thereof |
Non-Patent Citations (2)
| Title |
|---|
| CATHERINE DAUGA等: "Evolution of the gyrB gene and the molecular phylogeny of Enterobacteriaceae: a model molecule for molecular systematic studies", 《INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLOGY》 * |
| 安然等: "gyrB基因在细菌分类和检测中的应用", 《江西农业学报》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN112029830A (en) * | 2020-09-16 | 2020-12-04 | 上海上药第一生化药业有限公司 | Method for improving sensitivity of PCR (polymerase chain reaction) for identifying pig, cattle and sheep derived components in protein biochemical raw material crude product |
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