CN106048039A - Campylobacter jejuni rapid quantitative detection kit and using method - Google Patents
Campylobacter jejuni rapid quantitative detection kit and using method Download PDFInfo
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Abstract
The invention relates to the technology of biological detection, and aims to provide a campylobacter jejuni rapid quantitative detection kit. The kit comprises a plurality of freezing tubes, wherein the freezing tubes A are filled with freeze-dried powder, the freeze-dried powder comprises the following ingredients: a probe-method fluorescent quantitative PCR pre-mixed liquid reagent as well as two primers and a probe which are used for fluorescent quantitative detection; the five freezing tubes B are respectively filled with sterile double distilled water; the five freezing tubes C are respectively filled with TZ lysate; the total number of the freezing tubes D, E, F, G, H, I and J is seven, the freezing tubes D, E, F, G, H, I and J are respectively filled with campylobacter jejuni standard substance DNA freeze-dried powder with different concentrations, and the concentration of the campylobacter jejuni is decreased progressively by ten times. The campylobacter jejuni rapid quantitative detection kit is realized on the basis of design of primer probes on campylobacter jejuni mapA genes and optimization of detection conditions. By a campylobacter jejuni rapid quantitative detection method, the number of campylobacter jejuni in a quantitative sample can be detected within a short time, and the minimum lower limit of detection can reach 102 CFU/mL; the campylobacter jejuni rapid quantitative detection method has good specificity, sensitivity and accuracy, is convenient and speedy, and is easy to operate.
Description
Technical field
The invention belongs to Measurement for Biotechnique, particularly to the quick of food jejuni and detection by quantitative.It is intended to
Set up based on campylobacter jejuni fluorescence quantitative PCR Fast Detection Technique, i.e. campylobacter jejuni rapid quantitative detection reagent box and making
Use method.
Background technology
Campylobacter jejuni belongs to Campylobacter, is a kind of Gram-negative, microaerophilic spirilla, mainly causes people
Body acute enteritis, and the immune injury disease such as the reactive arthritis that occurs together, hepatitis, auspicious Te Shi disease and Guillain-Barre syndrome
Sick.1980, campylobacter enteritis was classified as one of most common infectious disease by World Health Organization (WHO) (WHO).2003, country of China
Campylobacter spp also will be classified as monitoring object by food origin disease monitoring net.
In many countries, cold chain has been supplied and has been exceeded the food of half and arrive consumer.Generally, this storage method quilt
Think it can is a kind of effective method reducing antibacterial, but the demand of fresh products is continuously increased, and creates in cold chain new
What risk was lived can not cultivation conditions antibacterial (VBNC).A large amount of pathogen (such as escherichia coli, Salmonella typhimurium, suddenly
Random coccus, helicobacter pylori and Listerella) VBNC state can be entered to tackle the differences such as cold, high ph-values and hyperosmosis
Ambient pressure.
Detection Campylobacter spp commonly uses traditional cultural method, and traditional method is the longest, generally takes 5～6 days and just can be detected
Result.Campylobacter jejuni can survive several weeks with " that lives can not cultivate " state in the environment, and traditional detection method can not have
These antibacterials detected to effect.Therefore, based on real-time quantitative PCR, detection based on hereditary basis and differentiate jejunum curved
The method of aspergillosis is particularly important to the detection of food-borne microorganism.
Summary of the invention
The technical problem to be solved in the present invention is, overcomes deficiency of the prior art, it is provided that a kind of campylobacter jejuni is quick
Immue quantitative detection reagent box and using method.
For solving technical problem, the solution of the present invention is:
A kind of campylobacter jejuni rapid quantitative detection reagent box is provided, including multiple cryopreservation tubes, the quantity of each cryopreservation tube and
Wherein content is specifically:
(1) cryopreservation tube of serial number A 1, equipped with lyophilized powder, its composition includes that sonde method quantitative fluorescent PCR premixed liquid tries
Agent, and it is used for two primers and the probe that fluorescent quantitation detects:
Primer CJ-F:5 '-TGTCAAGCACAACTATTCCTC-3 '
Primer CJ-R:5 '-TAAAAGAAGGGCAAAAGGT-3 '
Probe Taqman:5 '-FAM-ACCGCATTAAAATTCACATCGACA-TAMRA-3 '；
This lyophilized powder is prepared by following method:
Take sonde method quantitative fluorescent PCR premixed liquid reagent 1mL, described two each 0.9nM of primer, probe Taqman
0.2nM, lyophilizing after mixing, standby；
(2) cryopreservation tube of serial number B 5, is respectively provided with aseptic double-distilled water 2mL；
(3) cryopreservation tube of serial number C 5, is respectively provided with TZ lysate 1mL；
(4) serial number D～the cryopreservation tube of J, is respectively provided with the campylobacter jejuni standard substance DNA lyophilizing of variable concentrations by totally 7
Powder；Wherein, the cryopreservation tube of serial number D is jejunum campylobacter bacteria concentration be 108Bacterium solution DNA of CFU/mL, the cryopreservation tube of serial number E
In be the DNA of bacterium solution after bacterium solution in cryopreservation tube D dilutes ten times, the like.
Invention further provides the using method of described campylobacter jejuni rapid quantitative detection reagent box, including following
Step:
(1) dilution of cryopreservation tube A
Take the aseptic double-distilled water 1.8mL in cryopreservation tube B, add in cryopreservation tube A, resuspended, fully mix；
(2) sample DNA extracts
Take 1mL as the bacterial cultures of measuring samples, water sample, or buffer with the Sterile phosphate of 0.01M, pH 7.2
The flushing liquor on flushed solid sample surface；Measuring samples is centrifuged 2min with 12000rpm, abandons supernatant；With 500 μ L without
The resuspended precipitation of bacterium distilled water, 12000rpm is centrifuged 2min, abandons supernatant；Precipitation adds the TZ lysate in the cryopreservation tube C of 50 μ L
Fully mixing, 30min placed by-20 DEG C of refrigerators；Boiling water boiling 10min, ice bath 10min, 6000rpm be centrifuged 5min, collect supernatant,
DNA profiling as detected sample；
(3) standard substance dilution and sample-adding
Take the aseptic double-distilled water in cryopreservation tube B, add 50 μ L respectively in serial number D～J cryopreservation tube, concussion mixing；Take dilute
D～J cryopreservation tube after releasing contains the liquid of campylobacter jejuni standard substance DNA, and with the aseptic double-distilled water in 2 μ L cryopreservation tube B
As blank, being separately added in quantitative fluorescent PCR pipe, each concentration and blank arrange 2 repeating holes；Add the most respectively
Enter the diluent in 18 μ L step (1), mixing；
(4) measuring samples sample-adding
Take the DNA profiling of the detected sample extracted in 2 μ L step (2), add to quantitative fluorescent PCR pipe, each sample
Product arrange 2 repetitions；It is separately added into the diluent in 18 μ L step (1) again, mixing；
(5) fluorescence quantitative PCR detection
Take the quantitative fluorescent PCR pipe after sample-adding in step (3) and (4), put into and expand on quantitative real time PCR Instrument, bar
Part is: 95 DEG C of 5min；95 DEG C of 10sec, 56 DEG C of 30sec, 68 DEG C of 30sec gather signal, totally 40 circulations；
(6) Specification Curve of Increasing
Ct value meansigma methods and bacterial number according to standard substance draw standard curve；
(7) calculating of bacterial population in measuring samples
Sample amplification curve containing campylobacter jejuni is S-type, the Ct value of record sample, by the Ct value of measuring samples with empty
The standard curve that intestinal Campylobacter spp standard substance are drawn compares, the campylobacter jejuni quantity being calculated in measuring samples.
Compared with prior art, the beneficial effects of the present invention is:
1, the present invention is by campylobacter jejuni mapA gene design primed probe and the optimization of testing conditions, developing
A kind of campylobacter jejuni rapid quantitative detection reagent box.
2, the campylobacter jejuni fast quantitative measurement method for detecting that the present invention sets up can detect and in quantitative sample at short notice
Campylobacter jejuni quantity, lowest detection lower limit is up to 102CFU/mL, has good specificity, susceptiveness and accuracy, side
Just quick easily operation.
Accompanying drawing explanation
Fig. 1 is the standard curve drawn according to Ct value after standard substance bacterial number and certain amplification；
Fig. 2 is this test kit specific detection--campylobacter jejuni and the amplification curve of other antibacterials；
Fig. 3 is this test kit sensitivity Detection gradient concentration campylobacter jejuni；
Fig. 4 be this test kit sensitivity Detection repeat test；
Fig. 5 is that this test kit detects simulating pollution water sample；
Fig. 6 is that this test kit detects simulating pollution chicken meat sample.
In Fig. 3-6, open squares represents the sample clump count calculated according to fluorescent quantitation standard curve, black triangle
Represent some plate count results.
Detailed description of the invention
Campylobacter jejuni rapid quantitative detection reagent box in the present invention, by the cryopreservation tube of heterogeneity, can carry out 80 samples
The detection of product.
The campylobacter jejuni rapid quantitative detection reagent box of the present invention is contained difference by code name A, B, C, D, E, F, G, H, I and J
The cryopreservation tube composition of composition；1 cryopreservation tube A, equipped with sonde method quantitative fluorescent PCR premixed liquid reagent, (sequence is respectively primer
SEQ ID NO:1 and SEQ ID NO:2), probe (sequence is SEQ ID NO:3)；5 cryopreservation tube B, contain aseptic double steaming respectively
Water 2mL；5 cryopreservation tube C, contain TZ lysate 1mL respectively；Each one of cryopreservation tube D～J, contains the jejunum of variable concentrations respectively
Campylobacter spp standard substance DNA lyophilized powder, wherein, D pipe is 10 for concentration8DNA, the E pipe of CFU/mL campylobacter jejuni solution is for above-mentioned
The DNA of bacterium solution after bacterium solution dilution ten times, the like, J pipe dilutes bacterium solution DNA after 6 times for above-mentioned bacterium solution continuous ten times, the denseest
Degree is 102The DNA of CFU/mL campylobacter jejuni.
The sequence of above-mentioned primer and probe is shown in Table 1.
The primer of table 1 fluorescent quantitation and probe
The detecting step of campylobacter jejuni rapid quantitative detection reagent box is as follows:
(1) dilution of cryopreservation tube A: take the aseptic double-distilled water 1.8mL in cryopreservation tube B, adds to cryopreservation tube A, resuspended, the most mixed
Even；
(2) sample DNA extract: take 1mL as measuring samples bacterial cultures, water sample, or with Sterile phosphate buffer
Saline solution (0.01M pH 7.2) rinses the flushing liquor on solid sample surface；Measuring samples is centrifuged 2min with 12000rpm, abandons
Supernatant；By the 500 μ resuspended precipitations of L aseptic double-distilled water, 12000rpm is centrifuged 2min, abandons supernatant；Precipitation adds the cryopreservation tube of 50 μ L
TZ lysate in C fully mixes, and 30min placed by-20 DEG C of refrigerators；Boiling water boiling 10min, ice bath 10min, 6000rpm are centrifugal
5min, collects supernatant, as the DNA profiling of detected sample；
(3) standard substance dilution and sample-adding: take the aseptic double-distilled water in cryopreservation tube B, add 50 μ L respectively and freeze to serial number D～J
Deposit in pipe, concussion mixing；Take the liquid of (campylobacter jejuni standard substance DNA) in D～the J cryopreservation tube after dilution (with 2 μ L cryopreservation tubes
Aseptic double-distilled water in B is as blank), it is separately added in quantitative fluorescent PCR pipe, each concentration and blank arrange 2
Individual repeating hole；It is separately added into the diluent 18 μ L of step (1) again；Mixing；
(4) measuring samples sample-adding: take the sample DNA that 2 μ L step (2) extract, adds to quantitative fluorescent PCR pipe, each
Sample arranges 2 repetitions；It is separately added into the diluent 18 μ L of step (1) again；Mixing；
(5) detection: take the quantitative fluorescent PCR pipe after step (3) and (4) sample-adding, put into and expand on quantitative real time PCR Instrument
Increasing, condition is: 95 DEG C of 5min；95 DEG C of 10sec, 56 DEG C of 30sec, 68 DEG C of 30sec gather signal, totally 40 circulations；
(6) result judges: the sample amplification curve containing campylobacter jejuni is S-type, and the Ct value of record sample, according to standard
The Ct value meansigma methods of product and bacterial number (table 2) draw standard curve (Fig. 2), by the Ct value of sample and campylobacter jejuni standard substance
The standard curve drawn compares, and calculates the campylobacter jejuni quantity in sample.
Table 2 standard substance bacterial number
Specific embodiment:
One. cryopreservation tube forms
A pipe is lyophilized powder, containing sonde method quantitative fluorescent PCR premixed liquid reagent (AceQTM qPCR Probe Master
Mix, the product that Nanjing promise is only praised), primer and probe (shown in SEQ ID NO:1-3).It is stored in-20 DEG C.
B pipe, containing aseptic double-distilled water 2mL, is stored in-20 DEG C.
C pipe, containing TZ lysate 1mL, is stored in-20 DEG C.
Cryopreservation tube D～J is respectively provided with jejunum campylobacter bacteria concentration from 108C FU/mL to 102The campylobacter jejuni mark of CFU/mL
Quasi-product DNA lyophilized powder, is stored in-20 DEG C.
Two. sample-adding and detecting step explanation
Operate by the detecting step of aforementioned campylobacter jejuni rapid quantitative detection reagent box.
Below by test, the using effect of the present invention is proved and described.
1 materials and methods
1.1 bacterial strains and cultivation
Campylobacter jejuni reference strain is that zooprophylazis Institute for Medical Research of Zhejiang University separates preservation.Knot common in food
Intestinal Campylobacter spp (Campylobacter coli), escherichia coli (Enterobacteriaceae coli), monocyte hyperplasia Lee
This special bacterium (Listeria monocytogenes), Streptococcus suis (Streptococcus suis), Salmonella enteritidis
(Salmonella enteritidis), S. pullonum (Salmonella pullorum), vibrio parahaemolyticus
(Vibrio Parahemolyticus) also preserves for this institute.Campylobacter jejuni and Campylobacter Coli are inoculated in 5mL Bu Shi
Broth bouillon, 42 DEG C of micro-aerobic cultivation 48h.Other inoculation are in 5mL BHI culture medium, and 37 DEG C of shaken cultivation are overnight.
Prepared by 1.2 DNA profilings
Taking the PBS flushing liquor of 1mL bacterial cultures, water sample or solid sample, 12000rpm is centrifuged 2min, abandons supernatant.
Precipitating with the aqueous suspension in B pipe, 12000rpm is centrifuged 2min, abandons supernatant.Add the TZ lysate in the C pipe of 50 μ L the most mixed
Even ,-20 DEG C of refrigerators place 30min, boiling water boiling 10min, and ice bath 10min, 6000rpm are centrifuged 5min, and collection supernatant is template.
The preparation of 1.3 standard substance
Campylobacter jejuni culture 6000rpm is centrifuged 10min, adjusts bacterium solution OD by the resuspended precipitation of PBS620It is 0.12, this
Time campylobacter jejuni concentration be about 108CFU/mL, continuous ten times dilute 7 times, by the bacterium solution of 8 Concentraton gradient respectively at brother's rival
Put plate counting on sub-blood agar culture-medium, and respectively take 1mL and according to the method described above bacterial genomes DNA is extracted.
The foundation of 1.4 Real-time PCR system
1.4.1 design of primers
For specific gene mapA design pair of primers and a probe (table 1) in campylobacter jejuni, by Shanghai JaRa
Biological engineering company limited synthesizes.
1.4.2 Real-time PCR reaction system and condition
By Real-time PCR system and the optimization of condition, determine that reaction system is:
Reaction condition is:
1.4.3 specific test
By campylobacter jejuni, Campylobacter Coli, escherichia coli, Listeria monoeytogenes, Streptococcus suis, enteritis sand
Door Salmonella, S. pullonum and vibrio parahaemolyticus detect by fluorescent quantitation method after extracting DNA respectively, and use water
As negative control, investigate the specificity of fluorescent quantitative PCR detection method.
1.4.4 sensitivity and replica test
Jejunum campylobacter bacteria culture fluid is adjusted concentration to about 108CFU/mL, continuous ten times dilute 7 times, by 8 concentration ladders
The bacterium solution of degree point plate in Columbia Blood Agar culture medium counts and extracts DNA and carries out fluorescent quantitation amplification.According to glimmering
The result of light quantitative result and some plate counting determines the Monitoring lower-cut of the method.It is repeated once this test, checking this method
Repeatability.
1.4.5 analog sample detection
(1) simulating pollution chicken meat sample
The campylobacter jejuni culture of recovery is adjusted to about 108CFU/mL, ten times are diluted to 103CFU/mL.Each dilute
Degree of releasing takes the Carnis Gallus domesticus (Carnis Gallus domesticus is through Co60 14kGy irradiation) of 1mL inoculation 5g, after bacterium solution absorption completely, mixed with 20mLPBS vibration
Even, take diluent point plate and count and extract DNA and carry out fluorescent quantitation amplification.
(2) simulating pollution water sample
In view of water sample has other a fairly large number of bacterium, prepare mixed bacteria liquid to carry out fluorescent quantitation.Adjust large intestine bar
Bacteria concentration is about 109CFU/mL, jejunum campylobacter bacteria concentration is about 108CFU/mL, two kinds of bacterial suspensions put plate counting, then respectively
By two kinds of bacterial suspension equal-volume rear ten times of dilutions of mixing, it is fixed that each dilution bacterium solution test kit extracting DNA carries out fluorescence
Amount amplification.
The results contrast of the fluorescent quantitation result according to analog sample and some plate counting determines that the method is at actual sample
In feasibility.
1.6 actual sample detection sample-adding and detecting steps
1.6.1 take the aseptic double-distilled water 1.8mL in cryopreservation tube B, add to cryopreservation tube A, resuspended, fully mix；
1.6.2 take the bacterial cultures of 1mL, water sample or rinse with sterile phosphate buffer saline solution (0.01M pH 7.2) solid
The flushing liquor of body sample surfaces, 12000rpm is centrifuged 2min, abandons supernatant；With the 500 μ resuspended precipitations of L aseptic double-distilled water, 12000rpm
Centrifugal 2min, abandons supernatant；The TZ lysate added in precipitation in the cryopreservation tube C of 50 μ L fully mixes, and-20 DEG C of refrigerators are placed
30min；Boiling water boiling 10min, ice bath 10min, 6000rpm are centrifuged 5min, collect supernatant, as the DNA mould of detected sample
Plate；
1.6.3 take the aseptic double-distilled water in cryopreservation tube B, be separately added into serial number D～J cryopreservation tube 50 μ L, concussion mixing；Take
In D～J cryopreservation tube after dilution, the aseptic double-distilled water in the liquid of (campylobacter jejuni standard substance DNA) and cryopreservation tube B is (blank
Comparison) 2 μ L, it is separately added in quantitative fluorescent PCR pipe, each concentration and blank arrange 2 repeating holes；It is separately added into step again
The diluent 18 μ L of rapid 1.6.1；Mixing；
1.6.4 taking the sample DNA of 2 μ L step 1.6.2 extractings, add to quantitative fluorescent PCR pipe, each sample arranges 2
Individual repetition；It is separately added into the diluent 18 μ L of step 1.6.1 again；Mixing；
1.6.3 PCR pipe is closed the lid, put into and expand on quantitative real time PCR Instrument: 95 DEG C of 5min；(95 DEG C of 10sec,
56 DEG C of 30sec, 68 DEG C of 30sec gather signals) 40 circulations.
2 results
2.1 specific test
There is S type amplification curve through fluorescent quantitative PCR in campylobacter jejuni, and Campylobacter Coli, escherichia coli, monokaryon are thin
Born of the same parents' hyperplasia Listeria, Streptococcus suis, Salmonella enteritidis, S. pullonum and vibrio parahaemolyticus are without amplification curve
(Fig. 3), show that the method specificity is preferable.
2.2 sensitivity tests
Expanding the DNA sample of 8 dilution factor bacterium solution of campylobacter jejuni with fluorescent quantitation, be repeated twice, fluorescence is fixed
The results contrast of amount result and some plate counting is shown in Fig. 4 and Fig. 5.The result calculated by standard curve and actual point plate count results
Ratio is more consistent, and diversity is not notable (P ＞ 0.05).Double repeated experiment all shows that quantitative fluorescent PCR is to campylobacter jejuni
Low-detection lower limit is 102CFU/mL。
2.3 simulating pollution sample detection
By after Carnis Gallus domesticus simulating pollution and water sample simulating pollution gradient dilution extracting DNA carry out fluorescent quantitation detection (Fig. 5,
6), result shows, fluorescent quantitation standard curve the result calculated (P ＞ not notable with some plate count results diversity
0.05).Result show have Carnis Gallus domesticus or quantity bigger other bacterium interference time, the method still can detect that jejunum is curved specifically
Aspergillosis, Monitoring lower-cut is 102CFU/mL.Illustrate that this fluorescent quantitation method set up can apply to actual sample detection.
Result above all confirms the reliability of campylobacter jejuni rapid quantitative detection reagent box.
Claims (2)
1. a campylobacter jejuni rapid quantitative detection reagent box, it is characterised in that include multiple cryopreservation tube, the number of each cryopreservation tube
Measure with wherein content specifically:
(1) cryopreservation tube of serial number A 1, equipped with lyophilized powder, its composition includes sonde method quantitative fluorescent PCR premixed liquid reagent, with
And for fluorescent quantitation detection two primers and a probe:
Primer CJ-F:5 '-TGTCAAGCACAACTATTCCTC-3 '
Primer CJ-R:5 '-TAAAAGAAGGGCAAAAGGT-3 '
Probe Taqman:5 '-FAM-ACCGCATTAAAATTCACATCGACA-TAMRA-3 '；
This lyophilized powder is prepared by following method:
Take sonde method quantitative fluorescent PCR premixed liquid reagent 1mL, described two each 0.9nM of primer, probe Taqman 0.2nM, mixed
Even rear lyophilizing, standby；
(2) cryopreservation tube of serial number B 5, is respectively provided with aseptic double-distilled water 2mL；
(3) cryopreservation tube of serial number C 5, is respectively provided with TZ lysate 1mL；
(4) serial number D～the cryopreservation tube of J, is respectively provided with the campylobacter jejuni standard substance DNA lyophilized powder of variable concentrations by totally 7；
Wherein, the cryopreservation tube of serial number D is jejunum campylobacter bacteria concentration be 108Bacterium solution DNA of CFU/mL, in the cryopreservation tube of serial number E
It is the DNA of bacterium solution after the bacterium solution in cryopreservation tube D dilutes ten times, the like.
2. the using method of campylobacter jejuni rapid quantitative detection reagent box described in claim 1, it is characterised in that include following
Step:
(1) dilution of cryopreservation tube A
Take the aseptic double-distilled water 1.8mL in cryopreservation tube B, add in cryopreservation tube A, resuspended, fully mix；
(2) sample DNA extracts
Take 1mL as the bacterial cultures of measuring samples, water sample, or the sterile phosphate buffer saline with 0.01M, pH 7.2 is molten
Liquid rinses the flushing liquor on solid sample surface；Measuring samples is centrifuged 2min with 12000rpm, abandons supernatant；Aseptic double with 500 μ L
Steaming the resuspended precipitation of water, 12000rpm is centrifuged 2min, abandons supernatant；Precipitation adds the TZ lysate in the cryopreservation tube C of 50 μ L abundant
Mixing, 30min placed by-20 DEG C of refrigerators；Boiling water boiling 10min, ice bath 10min, 6000rpm be centrifuged 5min, collect supernatant, as
The DNA profiling of detected sample；
(3) standard substance dilution and sample-adding
Take the aseptic double-distilled water in cryopreservation tube B, add 50 μ L respectively in serial number D～J cryopreservation tube, concussion mixing；After taking dilution
D～J cryopreservation tube in containing the liquid of campylobacter jejuni standard substance DNA, and using the aseptic double-distilled water in 2 μ L cryopreservation tube B as
Blank, is separately added in quantitative fluorescent PCR pipe, and each concentration and blank arrange 2 repeating holes；It is separately added into 18 again
Diluent in μ L step (1), mixing；
(4) measuring samples sample-adding
Taking the DNA profiling of the detected sample extracted in 2 μ L step (2), add to quantitative fluorescent PCR pipe, each sample sets
Put 2 repetitions；It is separately added into the diluent in 18 μ L step (1) again, mixing；
(5) fluorescence quantitative PCR detection
Taking the quantitative fluorescent PCR pipe after sample-adding in step (3) and (4), put into and expand on quantitative real time PCR Instrument, condition is:
95℃5min；95 DEG C of 10sec, 56 DEG C of 30sec, 68 DEG C of 30sec gather signal, totally 40 circulations；
(6) Specification Curve of Increasing
Ct value meansigma methods and bacterial number according to standard substance draw standard curve；
(7) calculating of bacterial population in measuring samples
Sample amplification curve containing campylobacter jejuni is S-type, and the Ct value of record sample, by curved with jejunum for the Ct value of measuring samples
The standard curve that aspergillosis standard substance are drawn compares, the campylobacter jejuni quantity being calculated in measuring samples.
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Cited By (5)
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| CN106868160A (en) * | 2017-03-21 | 2017-06-20 | 杭州迪安生物技术有限公司 | Primer and its application of various diarrhoea pathogenic bacterias are detected simultaneously |
| CN107699629A (en) * | 2017-11-23 | 2018-02-16 | 深圳市草履虫生物科技有限公司 | Detect composite fluorescence PCR detection primer and probe, the kit of campylobacter jejuni, salmonella and Listeria monocytogenes |
| CN108424976A (en) * | 2018-02-08 | 2018-08-21 | 杭州富集生物科技有限公司 | Happen suddenly acute and severe infectious disease Emergent detection deposit kit and the method for inspection |
| CN109182567A (en) * | 2018-09-27 | 2019-01-11 | 军事科学院军事医学研究院环境医学与作业医学研究所 | A kind of method of real-time fluorescence quantitative PCR that is while detecting 12 kinds of pathogenic bacterias |
| CN116103363A (en) * | 2023-04-13 | 2023-05-12 | 江苏睿源生物技术有限公司 | Qualitative detection method for bacteria |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106868160A (en) * | 2017-03-21 | 2017-06-20 | 杭州迪安生物技术有限公司 | Primer and its application of various diarrhoea pathogenic bacterias are detected simultaneously |
| CN107699629A (en) * | 2017-11-23 | 2018-02-16 | 深圳市草履虫生物科技有限公司 | Detect composite fluorescence PCR detection primer and probe, the kit of campylobacter jejuni, salmonella and Listeria monocytogenes |
| CN108424976A (en) * | 2018-02-08 | 2018-08-21 | 杭州富集生物科技有限公司 | Happen suddenly acute and severe infectious disease Emergent detection deposit kit and the method for inspection |
| CN109182567A (en) * | 2018-09-27 | 2019-01-11 | 军事科学院军事医学研究院环境医学与作业医学研究所 | A kind of method of real-time fluorescence quantitative PCR that is while detecting 12 kinds of pathogenic bacterias |
| CN116103363A (en) * | 2023-04-13 | 2023-05-12 | 江苏睿源生物技术有限公司 | Qualitative detection method for bacteria |
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