CN108531658A - A kind of detection method of PEDV and TGEV primer sequences and duplex RT-PCR - Google Patents
A kind of detection method of PEDV and TGEV primer sequences and duplex RT-PCR Download PDFInfo
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Abstract
The invention discloses a kind of primer sequence of the double RT PCR of PEDV and TGEV detections, the primer sequence of the PEDV is sequence 1：ATGCTACAATTAGTGAATG, SEQ ID NO:1 sequence 2：TTATACGTCAATAACAGTAC, SEQ ID NO:2；The primer sequence sequence 3 of the TGEV：TAGACAAACTCGCTATCGCA, SEQ ID NO:3；Sequence 4:TTCTATCTGGTCGCCATCTT, SEQ ID NO:4；Double RT PCR detection methods, include the following steps：Step (1)：Total serum IgE is extracted from chitling road tissue sample；Step (2)：It is cDNA by the total serum IgE reverse transcription extracted in step (2), carries out PCR reactions using cDNA as template and using primer sequence, obtain PCR product；By PCR product after agarose gel electrophoresis, it is imaged and is taken pictures with the ultraviolet gel imaging system observations of GeL Doc TMXR.By the detection method of double RT PCR in the present invention, and change the response procedures of PCR, detection that can be fast and accurately and the method for telling PEDV and TGEV.
Description
Technical field
The present invention relates to biotechnology, more particularly to a kind of PEDV and TGEV primer sequences and double
The detection method of RT-PCR.
Background technology
Porcine epidemic diarrhea virus (porcine epidemic diarrhea virus, PEDV) and pig transmissible stomach and intestine
Scorching virus (transmissible gastroenteritis virus, TGEV) is the main pathogen for causing pig virus diarrhoea,
Stronger infectiousness is all had, especially most strong with the neurological susceptibility of suckling pig, lethality is high.Two kinds of viruses belong to coronavirus
Belong to member, very much like on pathogenic and epidemiology, using vomiting, diarrhea, dehydration as main feature, pathological change is similar,
It is clinically not easy to differentiate.Two kinds of viruses easily infect piglet, have similar clinical symptoms, epidemiology and pathological change, and
Usually there is mixed infection.Therefore, it relies solely on clinical observation to be difficult to be distinguished, Binding experiment room is needed to diagnose.
In recent years, PEDV is in rising trend in the incidence of multiple provinces and cities of China, be perplex pig breeding industry principal disease it
One.Since winter in 2010, PEDV is broken out in more than 10 a provinces and cities of China, and sick pig shows watery diarrhea, chordapsus, vomiting etc.
Classical symptom, the death rate is high, causes to inflict heavy losses on to China's pig breeding industry, causes the great attention of pig breeding industry.The master of current epidemic situation
Original of causing a disease is PEDV, however, also isolating TGEV out of some sick dead pig bodies.
Therefore, research it is a kind of efficiently, efficiently diagnostic techniques come distinguish PEDV and TGEV be those skilled in the art there is an urgent need for
It solves the problems, such as.
Invention content
In view of this, the present invention provides it is a kind of according to PEDV E gene orders and TGEV N genes separately design PEDV and
The primer of TGEV, by using duplex RT-PCR detection method can efficiently, efficiently tell PEDV and TGEV.
To achieve the goals above, the present invention adopts the following technical scheme that：A kind of duplex RT-PCR inspection of PEDV and TGEV
The primer sequence of survey,
The primer sequence of PEDV is sequence 1：ATGCTACAATTAGTGAATG, SEQ ID NO:1；Sequence 2：
TTATACGTCAATAACAGTAC, SEQ ID NO:2；
The primer sequence sequence 3 of TGEV：TAGACAAACTCGCTATCGCA, SEQ ID NO:3；Sequence 4:
TTCTATCTGGTCGCCATCTT, SEQ ID NO:4.
Preferably, the primer sequence of PEDV and TGEV is obtained according to PEDV E gene orders and the design of TGEV N genes.
The present invention also provides a kind of double PT-PCR detection methods for PEDV and TGEV, which is characterized in that including with
Lower step：
Step (1)：Total serum IgE is extracted from chitling road tissue sample；
Step (2)：It is cDNA by the total serum IgE reverse transcription extracted in step (2), primer sequence as template and is used using cDNA
PCR reactions are carried out, PCR product is obtained；By PCR product after agarose gel electrophoresis, with the ultraviolet gels of GeL Doc TMXR at
It is observed as system and is imaged and takes pictures.
Advantageous effect of the present invention：It is separately designed by using according to PEDV E gene orders and TGEV N genes in the present invention
The primer sequence of PEDV and TGEV by the detection method of double PT-PCR, and changes the response procedures of PCR, obtains one kind
Detection that can be fast and accurately and the method for telling PEDV and TGEV.
Preferably, PCR response procedures described in step (3) are 90 DEG C of -95 DEG C of pre-degeneration 2-3min；92 DEG C of -95 DEG C of denaturation
26-32s；48 DEG C of -62 DEG C of annealing 26-30s；70 DEG C of -73 DEG C of extension 1-3min；Totally 35 cycles；Last 70 DEG C -73 DEG C of extension 6-
10min；In 3 DEG C of -5 DEG C of preservations.
Preferably, PCR response procedures described in step (3) are 94 DEG C of pre-degeneration 3min；94 DEG C of denaturation 30s；58.1 DEG C moving back
Fiery 30s；72 DEG C of extension 1min；Totally 35 cycles；Last 72 DEG C of extensions 7min；In 4 DEG C of -5 DEG C of preservations.
Preferably, chitling road tissue sample described in step (2) is selected from swine excrement and small intestine.
Preferably, agarose gel described in step (3) is the agarose gel of 1%-3%, and the electrophoretic voltage is 110V, electricity
The swimming time is 30-35min.
It can be seen via above technical scheme that compared with prior art, the present disclosure provides a kind of pig epidemics
The detection method with transmissible gastroenteritis of swine duplex RT-PCR is rushed down, is reacted by PCR by using two kinds of primers in the present invention,
Efficiently, PEDV and TGEV are efficiently told.
Description of the drawings
In order to more clearly explain the embodiment of the invention or the technical proposal in the existing technology, to embodiment or will show below
There is attached drawing needed in technology description to be briefly described, it should be apparent that, the accompanying drawings in the following description is only this
The embodiment of invention for those of ordinary skill in the art without creative efforts, can also basis
The attached drawing of offer obtains other attached drawings.
Fig. 1 attached drawings are electrophoretogram of the PCR product through Ago-Gel in embodiment 1-8 in the present invention；
Fig. 2 attached drawings are electrophoretogram of the PCR product through Ago-Gel in comparative example 1 in the present invention.
Specific implementation mode
Following will be combined with the drawings in the embodiments of the present invention, and technical solution in the embodiment of the present invention carries out clear, complete
Site preparation describes, it is clear that described embodiments are only a part of the embodiments of the present invention, instead of all the embodiments.It is based on
Embodiment in the present invention, it is obtained by those of ordinary skill in the art without making creative efforts every other
Embodiment shall fall within the protection scope of the present invention.
The embodiment of the invention discloses a kind of double PT-PCR detection methods for PEDV and TGEV.
Embodiment 1
For the duplex RT-PCR detection method of PEDV and TGEV, include the following steps：
Step (1)：It is as follows that PEDV primer sequences are designed according to PEDV E gene orders：The primer sequence of PEDV is sequence 1：
ATGCTACAATTAGTGAATG, SEQ ID NO:1；Sequence 2：TTATACGTCAATAACAGTAC, SEQ ID NO:2；
It is as follows that TGEV primer sequences are designed according to TGEV N genes：The primer sequence of TGEV is sequence 3：
TAGACAAACTCGCTATCGCA, SEQ ID NO:3；Sequence 4:TTCTATCTGGTCGCCATCTT, SEQ ID NO:4；
Step (2)：Using RNA Miniprep Kits, total serum IgE is extracted from pig intestinal tissue；
Step (3)：It is cDNA by the total serum IgE reverse transcription extracted in step (2), carries out PCR reactions by template of cDNA, obtain
Obtain PCR product；PCR response procedures are 94 DEG C of pre-degeneration 3min；94 DEG C of denaturation 30s；48.0 DEG C of annealing 30s；72 DEG C of extension 1min；
Totally 35 cycles；Last 72 DEG C of extensions 7min；It is preserved at 4 DEG C.PCR product is passed through into agarose gel electrophoresis in the case where voltage is 110V
After 35min, it is imaged and is taken pictures with the ultraviolet gel imaging system observations of GeL Doc TMXR.
Embodiment 2
For the double PT-PCR detection methods of PEDV and TGEV, include the following steps：
Step (1)：It is as follows that PEDV primer sequences are designed according to PEDV E gene orders：The primer sequence of PEDV is sequence 1：
ATGCTACAATTAGTGAATG, SEQ ID NO:1；Sequence 2：TTATACGTCAATAACAGTAC, SEQ ID NO:2；
It is as follows that TGEV primer sequences are designed according to TGEV N genes：The primer sequence of TGEV is sequence 3：
TAGACAAACTCGCTATCGCA, SEQ ID NO:3；Sequence 4:TTCTATCTGGTCGCCATCTT, SEQ ID NO:4；
Step (2)：Using RNA Miniprep Kits, total serum IgE is extracted from pig intestinal tissue；
Step (3)：It is cDNA by the total serum IgE reverse transcription extracted in step (2), carries out PCR reactions by template of cDNA, obtain
Obtain PCR product；PCR response procedures are 94 DEG C of pre-degeneration 3min；94 DEG C of denaturation 30s；50.1 DEG C of annealing 30s；72 DEG C of extension 1min；
Totally 35 cycles；Last 72 DEG C of extensions 7min；It is preserved at 4 DEG C.PCR product is passed through into agarose gel electrophoresis in the case where voltage is 110V
After 35min, it is imaged and is taken pictures with the ultraviolet gel imaging system observations of GeL Doc TMXR.
Embodiment 3
For the duplex RT-PCR detection method of PEDV and TGEV, include the following steps：
Step (1)：It is as follows that PEDV primer sequences are designed according to PEDV E gene orders：The primer sequence of PEDV is sequence 1：
ATGCTACAATTAGTGAATG, SEQ ID NO:1；Sequence 2：TTATACGTCAATAACAGTAC, SEQ ID NO:2；
It is as follows that TGEV primer sequences are designed according to TGEV N genes：The primer sequence of TGEV is sequence 3：
TAGACAAACTCGCTATCGCA, SEQ ID NO:3；Sequence 4:TTCTATCTGGTCGCCATCTT, SEQ ID NO:4；
Step (2)：Using RNA Miniprep Kits, total serum IgE is extracted from pig intestinal tissue；
Step (3)：It is cDNA by the total serum IgE reverse transcription extracted in step (2), carries out PCR reactions by template of cDNA, obtain
Obtain PCR product；PCR response procedures are 94 DEG C of pre-degeneration 3min；94 DEG C of denaturation 30s；51.7 DEG C of annealing 30s；72 DEG C of extension 1min；
Totally 35 cycles；Last 72 DEG C of extensions 7min；It is preserved at 4 DEG C.PCR product is passed through into agarose gel electrophoresis in the case where voltage is 110V
After 35min, it is imaged and is taken pictures with the ultraviolet gel imaging system observations of GeL Doc TMXR.
Embodiment 4
For the duplex RT-PCR detection method of PEDV and TGEV, include the following steps：
Step (1)：It is as follows that PEDV primer sequences are designed according to PEDV E gene orders：The primer sequence of PEDV is sequence 1：
ATGCTACAATTAGTGAATG, SEQ ID NO:1；Sequence 2：TTATACGTCAATAACAGTAC, SEQ ID NO:2；
It is as follows that TGEV primer sequences are designed according to TGEV N genes：The primer sequence of TGEV is sequence 3：
TAGACAAACTCGCTATCGCA, SEQ ID NO:3；Sequence 4:TTCTATCTGGTCGCCATCTT, SEQ ID NO:4；
Step (2)：Using RNA Miniprep Kits, total serum IgE is extracted from pig intestinal tissue；
Step (3)：It is cDNA by the total serum IgE reverse transcription extracted in step (2), carries out PCR reactions by template of cDNA, obtain
Obtain PCR product；PCR response procedures are 94 DEG C of pre-degeneration 3min；94 DEG C of denaturation 30s；53.7 DEG C of annealing 30s；72 DEG C of extension 1min；
Totally 35 cycles；Last 72 DEG C of extensions 7min；It is preserved at 4 DEG C.PCR product is passed through into agarose gel electrophoresis in the case where voltage is 110V
After 35min, it is imaged and is taken pictures with the ultraviolet gel imaging system observations of GeL Doc TMXR.
Embodiment 5
For the duplex RT-PCR detection method of PEDV and TGEV, include the following steps：
Step (1)：It is as follows that PEDV primer sequences are designed according to PEDV E gene orders：The primer sequence of PEDV is sequence 1：
ATGCTACAATTAGTGAATG, SEQ ID NO:1；Sequence 2：TTATACGTCAATAACAGTAC, SEQ ID NO:2；
It is as follows that TGEV primer sequences are designed according to TGEV N genes：The primer sequence sequence 3 of TGEV：
TAGACAAACTCGCTATCGCA, SEQ ID NO:3；Sequence 4:TTCTATCTGGTCGCCATCTT, SEQ ID NO:4；
Step (2)：Using RNA Miniprep Kits, total serum IgE is extracted from pig intestinal tissue；
Step (3)：It is cDNA by the total serum IgE reverse transcription extracted in step (2), carries out PCR reactions by template of cDNA, obtain
Obtain PCR product；PCR response procedures are 94 DEG C of pre-degeneration 3min；94 DEG C of denaturation 30s；56.0 DEG C of annealing 30s；72 DEG C of extension 1min；
Totally 35 cycles；Last 72 DEG C of extensions 7min；It is preserved at 4 DEG C.PCR product is passed through into agarose gel electrophoresis in the case where voltage is 110V
After 35min, it is imaged and is taken pictures with the ultraviolet gel imaging system observations of GeL Doc TMXR.
Embodiment 6
For the duplex RT-PCR detection method of PEDV and TGEV, include the following steps：
Step (1)：It is as follows that PEDV primer sequences are designed according to PEDV E gene orders：The primer sequence of PEDV is sequence 1：
ATGCTACAATTAGTGAATG, SEQ ID NO:1；Sequence 2：TTATACGTCAATAACAGTAC, SEQ ID NO:2；
It is as follows that TGEV virus primer sequences are designed according to TGEV N genes：The primer sequence of TGEV viruses is sequence 3：
TAGACAAACTCGCTATCGCA, SEQ ID NO:3；Sequence 4:TTCTATCTGGTCGCCATCTT, SEQ ID NO:4；
Step (2)：Using RNA Miniprep Kits, total serum IgE is extracted from pig intestinal tissue；
Step (3)：It is cDNA by the total serum IgE reverse transcription extracted in step (2), carries out PCR reactions by template of cDNA, obtain
Obtain PCR product；PCR response procedures are 94 DEG C of pre-degeneration 3min；94 DEG C of denaturation 30s；58.1 DEG C of annealing 30s；72 DEG C of extension 1min；
Totally 35 cycles；Last 72 DEG C of extensions 7min；It is preserved at 4 DEG C.PCR product is passed through into agarose gel electrophoresis in the case where voltage is 110V
After 35min, it is imaged and is taken pictures with the ultraviolet gel imaging system observations of GeL Doc TMXR.
Embodiment 7
For the duplex RT-PCR detection method of PEDV and TGEV, include the following steps：
Step (1)：It is as follows that PEDV primer sequences are designed according to PEDV E gene orders：The primer sequence of PEDV is sequence 1：
ATGCTACAATTAGTGAATG, SEQ ID NO:1；Sequence 2：TTATACGTCAATAACAGTAC, SEQ ID NO:2；
It is as follows that TGEV primer sequences are designed according to TGEV N genes：The primer sequence of TGEV is sequence 3：
TAGACAAACTCGCTATCGCA, SEQ ID NO:3；Sequence 4:TTCTATCTGGTCGCCATCTT, SEQ ID NO:4；
Step (2)：Using RNA Miniprep Kits, total serum IgE is extracted from pig intestinal tissue；
Step (3)：It is cDNA by the total serum IgE reverse transcription extracted in step (2), carries out PCR reactions by template of cDNA, obtain
Obtain PCR product；PCR response procedures are 94 DEG C of pre-degeneration 3min；94 DEG C of denaturation 30s；60.9 DEG C of annealing 30s；72 DEG C of extension 1min；
Totally 35 cycles；Last 72 DEG C of extensions 7min；It is preserved at 4 DEG C.PCR product is passed through into agarose gel electrophoresis in the case where voltage is 110V
After 35min, it is imaged and is taken pictures with the ultraviolet gel imaging system observations of GeL Doc TMXR.
Embodiment 8
For the duplex RT-PCR detection method of PEDV and TGEV, include the following steps：
Step (1)：It is as follows that PEDV primer sequences are designed according to PEDV E gene orders：The primer sequence of PEDV is sequence 1：
ATGCTACAATTAGTGAATG, SEQ ID NO:1；Sequence 2：TTATACGTCAATAACAGTAC, SEQ ID NO:2；
It is as follows that TGEV virus primer sequences are designed according to TGEV N genes：The primer sequence of TGEV is sequence 3：
TAGACAAACTCGCTATCGCA, SEQ ID NO:3；Sequence 4:TTCTATCTGGTCGCCATCTT, SEQ ID NO:4；
Step (2)：Using RNA Miniprep Kits, total serum IgE is extracted from pig intestinal tissue；
Step (3)：It is cDNA by the total serum IgE reverse transcription extracted in step (2), carries out PCR reactions by template of cDNA, obtain
Obtain PCR product；PCR response procedures are 94 DEG C of pre-degeneration 3min；94 DEG C of denaturation 30s；62.0 DEG C of annealing 30s；72 DEG C of extension 1min；
Totally 35 cycles；Last 72 DEG C of extensions 7min；It is preserved at 4 DEG C.PCR product is passed through into agarose gel electrophoresis in the case where voltage is 110V
After 35min, it is imaged and is taken pictures with the ultraviolet gel imaging system observations of GeL Doc TMXR.
Attached drawing 2 is the electrophoretogram of embodiment 1-8, from attached drawing 2 it can be seen that in the range of 53.7 DEG C -62.0 DEG C, energy
Enough preferably to detect two kinds of viruses, band when from electrophoresis it can be seen from the figure that at 58.1 DEG C is most clearly, therefore, most preferably
Annealing temperature is 58.1 DEG C.
Comparative example 1
Step (1)：It is as follows that PEDV primer sequences are designed according to PEDV E gene orders：The primer sequence of PEDV is sequence 1：
ATGCTACAATTAGTGAATG, SEQ ID NO:1；Sequence 2：TTATACGTCAATAACAGTAC, SEQ ID NO:2；
It is as follows that TGEV primer sequences are designed according to TGEV N genes：The primer sequence of TGEV is sequence 3：
TAGACAAACTCGCTATCGCA, SEQ ID NO:3；Sequence 4:TTCTATCTGGTCGCCATCTT, SEQ ID NO:4；
Step (2)：Using RNA Miniprep Kits, from the pig intestinal tissue of infection TGEV, PEDV, HCV, PRV and RV
Middle extraction total serum IgE；
Step (3)：It is cDNA by the total serum IgE reverse transcription extracted in step (2), carries out PCR reactions by template of cDNA, obtain
Obtain PCR product；PCR response procedures are 94 DEG C of pre-degeneration 3min；94 DEG C of denaturation 30s；58.1 DEG C of annealing 30s；72 DEG C of extension 1min；
Totally 35 cycles；Last 72 DEG C of extensions 7min；It is preserved at 4 DEG C.PCR product is passed through into agarose gel electrophoresis in the case where voltage is 110V
After 35min, it is imaged and is taken pictures with the ultraviolet gel imaging system observations of GeL Doc TMXR.
Attached drawing 2 is the electrophoretogram of comparative example 1, and 1-5 is respectively TGEV, PEDV, HCV, PRV and RV, as seen from the figure, two kinds
Primer TGEV that can be detected and PEDV, and HCV, PRV and RV in comparative example 1 can not be examined in the accompanying drawings without band
Measure its virus.
Pattern detection
The 135 parts of doubtful intestine of young pigs tissue samples with PEDV and TGEV that will be collected in January, 2014 in December, 2016
Piece total serum IgE, be respectively adopted conventional RT-PCR be the duplex PCR detection method in control group and embodiment 6 be test group detection
PEDV and TGEV.As a result such as table 1
Table 1
By in table 1, it can be deduced that the sample detected using embodiment 6 optimal in the present invention is more accurate, and uses
Conventional method can not detect the virus of PEDV and TGEV mixing.Show the detection method high sensitivity in the present invention.
Each embodiment is described by the way of progressive in this specification, the highlights of each of the examples are with other
The difference of embodiment, just to refer each other for identical similar portion between each embodiment.For device disclosed in embodiment
For, since it is corresponded to the methods disclosed in the examples, so description is fairly simple, related place is said referring to method part
It is bright.
The foregoing description of the disclosed embodiments enables those skilled in the art to implement or use the present invention.
Various modifications to these embodiments will be apparent to those skilled in the art, as defined herein
General Principle can be realized in other embodiments without departing from the spirit or scope of the present invention.Therefore, of the invention
It is not intended to be limited to the embodiments shown herein, and is to fit to and the principles and novel features disclosed herein phase one
The widest range caused.
Sequence table
<110>Longyan School
<120>A kind of detection method of PEDV and TGEV primer sequences and duplex RT-PCR
<160> 4
<170> SIPOSequenceListing 1.0
<210> 1
<211> 19
<212> DNA
<213> porcine epidemic diarrhea virus
<400> 1
atgctacaat tagtgaatg 19
<210> 2
<211> 20
<212> DNA
<213> porcine epidemic diarrhea virus
<400> 2
ttatacgtca ataacagtac 20
<210> 3
<211> 20
<212> DNA
<213> Porcine transmissible gastroenteritis virus
<400> 3
tagacaaact cgctatcgca 20
<210> 4
<211> 20
<212> DNA
<213> Porcine transmissible gastroenteritis virus
<400> 4
ttctatctgg tcgccatctt 20
Claims (7)
1. a kind of primer sequence of the duplex RT-PCR detection of PEDV and TGEV, which is characterized in that
The primer sequence of the PEDV is sequence 1：ATGCTACAATTAGTGAATG, SEQ ID NO:1；Sequence 2：
TTATACGTCAATAACAGTAC, SEQ ID NO:2；
The primer sequence of the TGEV is sequence 3：TAGACAAACTCGCTATCGCA, SEQ ID NO:3；Sequence 4:
TTCTATCTGGTCGCCATCTT, SEQ ID NO:4.
2. the primer sequence of the duplex RT-PCR detection of PEDV and TGEV according to claim 1, which is characterized in that described
The primer sequence of PEDV and TGEV is obtained according to PEDV E gene orders and the design of TGEV N genes.
3. a kind of duplex RT-PCR detection method for PEDV and TGEV, which is characterized in that include the following steps：
Step (1)：Total serum IgE is extracted from chitling road tissue sample；
Step (2)：It is cDNA by the total serum IgE reverse transcription extracted in step (2), using cDNA as template and using claim 1-2
Any one of obtain primer sequence carry out PCR reactions, obtain PCR product；By PCR product after agarose gel electrophoresis, use
The ultraviolet gel imaging system observations of GeL Doc TMXR are imaged and take pictures.
4. a kind of duplex RT-PCR detection method for PEDV and TGEV according to claim 3, which is characterized in that step
Suddenly PCR response procedures described in (3) are 90 DEG C of -95 DEG C of pre-degeneration 2-3min；92 DEG C of -95 DEG C of denaturation 26-32s；48 DEG C -62 DEG C are moved back
Fiery 26-30s；70 DEG C of -73 DEG C of extension 1-3min；Totally 35 cycles；Last 70 DEG C -73 DEG C of extension 6-10min；In 3 DEG C of -5 DEG C of guarantors
It deposits.
5. a kind of duplex RT-PCR detection method for PEDV and TGEV according to claim 3 or 4, feature exist
In PCR response procedures described in step (3) are 94 DEG C of pre-degeneration 3min；94 DEG C of denaturation 30s；58.1 DEG C of annealing 30s；72 DEG C are prolonged
Stretch 1min；Totally 35 cycles；Last 72 DEG C of extensions 7min；In 4 DEG C of -5 DEG C of preservations.
6. a kind of duplex RT-PCR detection method for PEDV and TGEV according to claim 3 or 4, feature exist
In chitling road tissue sample described in step (2) is selected from swine excrement and small intestine.
7. a kind of duplex RT-PCR detection method for PEDV and TGEV according to claim 3 or 4, feature exist
In agarose gel described in step (3) is the agarose gel of 1%-3%, and the electrophoretic voltage is 110V, electrophoresis time 30-
35min。
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| CN201810437946.2A CN108531658A (en) | 2018-05-09 | 2018-05-09 | A kind of detection method of PEDV and TGEV primer sequences and duplex RT-PCR |
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| CN201810437946.2A CN108531658A (en) | 2018-05-09 | 2018-05-09 | A kind of detection method of PEDV and TGEV primer sequences and duplex RT-PCR |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN109097499A (en) * | 2018-09-19 | 2018-12-28 | 浙江农林大学 | It is a kind of for detecting the dual fluorescence quantification RT-PCR detection kit of PEDV-TGEV nucleic acid |
| CN109825649A (en) * | 2019-04-09 | 2019-05-31 | 广西大学 | Porcine epidemic diarrhea virus G1/G2 type RT-PCR diagnostic primers group and its diagnostic kit |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109097499A (en) * | 2018-09-19 | 2018-12-28 | 浙江农林大学 | It is a kind of for detecting the dual fluorescence quantification RT-PCR detection kit of PEDV-TGEV nucleic acid |
| CN109825649A (en) * | 2019-04-09 | 2019-05-31 | 广西大学 | Porcine epidemic diarrhea virus G1/G2 type RT-PCR diagnostic primers group and its diagnostic kit |
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Application publication date: 20180914 |