CN107326082B - Universal primer of human cestode ep45 gene and identification method thereof - Google Patents
Universal primer of human cestode ep45 gene and identification method thereof Download PDFInfo
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Abstract
The invention relates to the technical field of molecular biology, and discloses universal primers and nucleotide sequences of three human cestode ep45 genes and an identification method thereof. The primer comprises an upstream primer shown as SEQ ID NO. 1 and a downstream primer shown as SEQ ID NO. 2; wherein, SEQ ID NO: 1: 5'-AGAGACAAAGGAAAGGCAGCTG-3', respectively; 2, SEQ ID NO: 5'-GGGGCTCAGGCACCTTGAT-3', respectively; according to the embodiment of the invention, universal primers of three human cestode ep45 genes are designed, RT-PCR amplification is carried out to obtain an ep45 gene fragment of the cestode, then enzyme digestion treatment is carried out by using EcoR I and/or Aar I according to the difference of restriction enzyme sites among the three ep45, and interspecies identification is carried out on the three human cestode according to the difference (1/2/3 bands) of the electrophoretic appearance map of the enzyme digestion product, so that the time for sequencing identification of the PCR product is greatly shortened, and the judgment result according to the enzyme digestion map is more accurate and reliable.
Description
Technical Field
The invention relates to the field of molecular biology, in particular to universal primers and nucleotide sequences of three human cestode ep45 genes and an identification method thereof.
Background
Taenia solium, taenia asialica and taenia bovis seu Bubali are important zoonotic parasites, which are parasitic in human small intestine and cause taeniasis and even cysticercosis in human. Therefore, the identification of the species has important significance in the related research and the comprehensive prevention and treatment of the taeniasis/cysticercosis. The three tapeworms parasitizing in human intestinal tracts are very similar in shape, and the most important difference between the three is whether the knob is provided with a tip protrusion or a small hook. Thus, the conventional methods are distinguished by microscopic observation of the head segment after tabletting. Unfortunately, the head segment of the worm body is usually lost, and the worm body is difficult to identify by simply relying on the segment or the worm egg, and usually needs to be identified by a molecular biological method. The most common identification method is to amplify 18s or mitochondrial gene sequences of three polypide by PCR respectively, then carry out sequencing analysis and identify according to the difference of target sequences. However, this method has the disadvantage that sequencing is time consuming and that accurate interspecific identification is not possible when the sequence similarity is close (hybrids).
Disclosure of Invention
The embodiment of the invention aims to provide three methods for identifying human tapeworms, which not only overcome the defects of the traditional microscopic examination, but also carry out single-gene PCR amplification and enzyme digestion identification through a universal primer, can realize the molecular identification of the tapeworms without gene sequencing, greatly accelerate the speed of the tapeworm identification, and simultaneously enable the identification result of the tapeworms to be more accurate and reliable through a method for comparing enzyme digestion maps.
In order to solve the technical problems, the embodiment of the invention provides three universal primers of human tapeworm ep45 gene, which comprise an upstream primer shown as SEQ ID NO. 1 and a downstream primer shown as SEQ ID NO. 2; wherein, SEQ ID NO: 1: 5'-AGAGACAAAGGAAAGGCAGCTG-3', respectively; 2, SEQ ID NO: 5'-GGGGCTCAGGCACCTTGAT-3' are provided.
The embodiment of the invention also provides nucleotide sequences of three human tapeworms ep45, wherein the three human tapeworms are taenia solium, taenia bovis and taenia asiana; the nucleotide sequence of ep45 of taenia solium is shown in SEQ ID NO 3; the nucleotide sequence of ep45 of Taenia bovis is set forth in SEQ ID NO 4; the nucleotide sequence of ep45 from Taenia Asiatica is shown in SEQ ID NO 5.
The embodiment of the invention also provides three identification methods of the human tapeworm, which comprise the following steps:
(1) extracting three kinds of total RNA of the human tapeworm to be identified, and carrying out reverse transcription on the total RNA by the universal primer to synthesize cDNA of the human tapeworm to be identified;
(2) carrying out ep45 gene PCR amplification by taking the human tapeworm cDNA to be identified as a template;
(3) carrying out agarose gel electrophoresis on the PCR amplification product, and then carrying out gel recovery; obtaining an ep45 glue recovery product;
(4) carrying out enzyme digestion treatment on the recovered product of the ep45 gel by using EcoRI and/or AarI restriction endonuclease;
(5) and (4) carrying out agarose gel electrophoresis on the enzyme digestion product, and identifying the human tapeworm according to the enzyme digestion map.
Compared with the prior art, the embodiment of the invention can obtain the ep45 gene fragment of three human tapeworms by RT-PCR amplification by designing the universal primer of the ep45 gene, then carry out enzyme digestion treatment by EcoRI and/or AarI according to the difference of restriction enzyme sites among the three, and carry out interspecies identification on the human tapeworms according to the difference (1 strip/2 strips/3 strips) of the appearance map of the electrophoresis of the enzyme digestion product, thereby greatly shortening the identification time of the human tapeworms, and the judgment result of the enzyme digestion map is more reliable than that of sequence comparison.
Preferably, the conditions for PCR amplification of the ep45 gene are: pre-denaturation at 95 ℃ for 5 min; denaturation at 94 ℃ for 1min, annealing at 60 ℃ for 1min, and extension at 72 ℃ for 90s for 30 cycles; after the end of the last cycle, the extension was carried out for 10min at 72 ℃.
Preferably, the reaction system for PCR amplification of the ep45 gene is as follows: rTaq texture: 25.0 μ L; 1, SEQ ID NO: 1.0 μ L; 2, SEQ ID NO: 1.0 μ L; and (3) identifying human tapeworm cDNA: 1.0 μ L; DEPC H2O: 22.0. mu.L.
Preferably, the gel recovery is performed using a DNA gel in step 3.
Preferably, the total RNA of the human tapeworm to be identified is extracted from a tapeworm segment or an egg.
Preferably, the enzyme cutting system of the ep45 gene is as follows:
the ep45 glue recovers 5 μ L of the product;
EcoRI or AarI 1. mu.L;
Buffer 2μL；
11.5 μ L deionized water.
Drawings
FIG. 1 is a cleavage map of a single cleavage of a recovered product of a gel according to a second embodiment of the present invention.
1. TS control 12; 2. 11, TSA control; 3. TAS control 10; 4, carrying out enzyme digestion on TSep45 EcoRI; 5: carrying out enzyme digestion on the TSAep45 EcoRI; 6, TASep45 EcoRI enzyme cutting result; 7, TSep45AarI enzyme digestion result; 8, performing enzyme digestion on the TSAep45 AarI; 9, TASep45 AarI cleavage result.
Detailed Description
The following examples are given to facilitate a better understanding of the invention, but do not limit the invention. The experimental procedures in the following examples are conventional unless otherwise specified. The test materials used in the following examples were purchased from a conventional biochemical reagent store unless otherwise specified.
The first embodiment of the invention relates to universal primers of three human cestode ep45 genes, which comprise an upstream primer shown as SEQ ID NO. 1 and a downstream primer shown as SEQ ID NO. 2; wherein the content of the first and second substances,
SEQ ID NO:1：5’-AGAGACAAAGGAAAGGCAGCTG-3’
SEQ ID NO:2：5’-GGGGCTCAGGCACCTTGAT-3’。
the second embodiment of the invention relates to a method for identifying three human tapeworms, which comprises the following steps:
(1) sequence analysis of three human cestodes ep45
Downloading the nucleotide sequences of ep45 genes of three Tapeworms (TS), Taenia bovis (TSA) and Taenia Asiatica (TAS) from a Taenia genomic database of Lanzhou veterinary institute, and comparing the nucleotide sequences of ep45 of the three tapeworms by Blast analysis, wherein the nucleotide sequence of ep45 of the Taenia suis is shown in SEQ ID NO. 3; the nucleotide sequence of ep45 of Taenia bovis is set forth in SEQ ID NO 4; the nucleotide sequence of ep45 from Taenia Asiatica is shown in SEQ ID NO 5;
(2) design of universal primers for three human cestodes ep45
Primers are designed by Primer Premier 5.0 software, the upstream Primer (ep45UPM-F) is at 18-40 sites of ORF, the downstream Primer (ep45UPM-R) is at 1305-1329 sites, the length of amplified fragments is 1306bp,
ep45UPM-F (i.e., SEQ ID NO:1):
5’-AGAGACAAAGGAAAGGCAGCTG-3’；
ep45UPM-R (i.e., SEQ ID NO:2):
5’-GGGGCTCAGGCACCTTGAT-3’；
(3) PCR amplification of ep45 Gene
Collecting about 30mg of human tapeworm section or worm egg to be detected, extracting total RNA by Trizol, then carrying out reverse transcription by using ep45UPM-R (SEQ ID NO:2) as a primer, synthesizing a cDNA first chain, and further carrying out PCR amplification by using cDNA as a template, wherein the conditions of the PCR amplification are as follows: pre-denaturation at 95 ℃ for 5 min; denaturation at 94 ℃ for 1min, annealing at 60 ℃ for 1min, and extension at 72 ℃ for 90s for 30 cycles; after the end of the last cycle, the extension was carried out for 10min at 72 ℃.
The reaction system for the PCR amplification of the ep45 gene is as follows:
rTaq texture (available from TaKaRa Co.): 25.0 μ L;
SEQ ID NO:1：1.0μL；
SEQ ID NO:2：1.0μL；
and (3) identifying human tapeworm cDNA: 1.0 μ L;
DEPC H2O 22.0μL。
(4) recovery and purification of PCR product of ep45 gene
The PCR product was observed by 1% agarose gel electrophoresis, and the gel strip of the target fragment was cut off and the target fragment was recovered by a DNA gel recovery kit.
(5) Digestion reaction of PCR product
Carrying out enzyme digestion treatment on the glue recovery product in the step 4 by using EcoRI and/or AarI restriction endonuclease; wherein the content of the first and second substances,
an enzyme digestion reaction system:
the EcoRI and/or AarI restriction enzymes are used in this embodiment for the following reasons: restriction enzyme sites of the three ep45 sequences were analyzed using DNAstar-seqBuilder restriction site software. Because the nucleotide sequence of the ep45 of the taenia solium has EcoRI and AarI 2 restriction sites, the nucleotide sequence of the ep45 of the taenia solium only has one EcoRI restriction site, and the nucleotide sequence of the ep45 of the taenia solium does not have the 2 restriction sites. Therefore, EcoRI and/or AarI restriction enzymes can be used for carrying out single-enzyme digestion or double-enzyme digestion reaction on the ep45 PCR product of a sample to be identified, then, agarose gel electrophoresis is carried out on the digestion product, and the three human tapeworms are identified according to the number or size (digestion pattern) of the bands.
(6) Band identification
If the gel recovery product in the step 4 is subjected to EcoRI single enzyme digestion treatment, 2 bands (766bp and 540bp) appear, the sample to be identified is Taenia Solium (TS), and if 1 band (1306bp) appears, the sample is Taenia Solium (TSA) or Taenia Asiana (TAS);
when the recovered product of the gel in step 4 is digested with AarI, it is TSA (697bp, 609bp) if 2 bands appear, and TAS if 1 band (1306bp) appears, as shown in FIG. 1.
If 3 bands (609bp, 540bp and 157bp) appear after the gel recovery product in the step 4 is subjected to double enzyme digestion treatment by EcoRI and AarI (shown in Table 1), the sample to be detected is Taenia Solium (TS);
if 2 bands (697bp, 609bp) appear after the gel recovery product in the step 4 is subjected to EcoRI and AarI double enzyme digestion treatment, the sample to be detected is Taenia bovis Hendel (TSA);
if 1 band (1306bp) appears after the gel recovery product in the step 4 is subjected to EcoRI and AarI double enzyme digestion treatment, the sample to be detected is the Taenia Asiatica (TAS) (not shown in the figure). Therefore, the glue recovery product in the step 4 can be processed by EcoRI or AarI alone or in combination, and the three human tapeworms can be subjected to differential diagnosis.
| Human tapeworm | EcoRI | AarI | EcoRI and AarI |
| TS | 766bp,540bp | 609bp,697bp | 609bp,540bp,157bp |
| TSA | 1306bp | 609bp,697bp | 609bp,697bp |
| TAS | 1306bp | 1306bp | 1306bp |
TABLE 1 enzyme digestion maps of three human cestode ep45 gene fragments
Compared with the prior art, in the research of the serpin inhibitor (serpins) gene family, the (ep45) gene which is one of serpins is found to be very conserved in taenia solium (taeniasisolium), taenia asialica (taeniasiastatica) and taenia bovis (taeniasisiaginata), and the consistency (Identity) of the three genes on the nucleic acid level is up to 96.4%, and the similarity (similarity) is up to 99.6%. Therefore, in the embodiment, on the basis of cloning and sequence analysis of the ep45 genes of the three human tapeworms, the ep45 gene segments (1306bp) of the three human tapeworms can be obtained by selecting a very conservative sequence to design a universal primer of the ep45 gene and performing RT-PCR. Then according to the difference of restriction enzyme sites among the three, carrying out enzyme digestion treatment by using EcoRI or AarI, and according to the difference (1/2/3 bands) of the appearance map of the enzyme digestion product electrophoresis, carrying out interspecies identification on three human tapeworms, greatly shortening the time of traditional molecular identification, and the judgment result of the enzyme digestion map is more reliable than that of sequence comparison.
It will be understood by those of ordinary skill in the art that the foregoing embodiments are specific examples for carrying out the invention, and that various changes in form and details may be made therein without departing from the spirit and scope of the invention in practice.
Sequence listing
<110> Lanzhou veterinary research institute of Chinese academy of agricultural sciences
<120> universal primer of human cestode ep45 gene and identification method thereof
<130> 2017
<160> 5
<170> SIPOSequenceListing 1.0
<210> 1
<211> 22
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 1
agagacaaag gaaaggcagc tg 22
<210> 2
<211> 19
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 2
ggggctcagg caccttgat 19
<210> 3
<211> 1350
<212> DNA
<213> Taenia solium (Taenia solium)
<400> 3
atgatttctc tacgtatgag agacaaagga aaggcagctg catctgagcc tgagagaccc 60
aagtctactt tttcgaaaag cgtcaaagtc aatccggtcg actatctgaa gcacagggac 120
ttcaacaaca aatattgctt cggtgttcat gccattaatg acatcaccaa gaagtcgggt 180
cactcgccaa caacgatacg cttcctcctg acagtgttgg tgggctcgaa ggcggcacgg 240
ggcactagtg cggaccagat cacacaggca ctcagcacca cgaactcacg tgacgtgggc 300
accgaatgca ccagcctcgt ggacagcgcg cttgacgagt gggctctgct ctctactgcg 360
ggtctaagcg atataccact cgatcgcatt gaggaagggc gcctctgtcg ctttcagtct 420
gctatcttcg tccccatgga tgatctcacc ttcaaggccg agttccagtg gttgatcacc 480
aaccgcatgg gtgtcgtctg gactcagact tcgagaaagg atttcgccca tgctcagaag 540
tggttatcca aagtgtcaaa aggtttattc actcaccact ttccaaaaca atgtgccaac 600
tctattgtac ttgccacagc attacaattt aaaggcaggt ggactcaacc tttggaactc 660
tacggtagtt ccaagggatc attcgaagtg tcaccccact cgaagatcga tgttcctatg 720
ctaaaaattt caacaaaggt tatttactat aaggatacca aaaaaggctt ccatttggtt 780
ggaattccac ttaaggatgc gcggttcgca atcgcatttc tattaccctt ggtgccgcac 840
aagttcgtag aggtggagca tcgctttaca gagggtttga actacggtct cttcaacagc 900
tcgcgactgc atttctgcag catgcacgtg gtcattccta tgttccgagt gcaaacggag 960
gtggacctct gtaaggcctt gcccttcctc ggcatgagca atccattcga ttctgatcgg 1020
gcagacttct ccggtatcag cgatgtagag aatctgcata tcaacagcgg caaggagtcg 1080
gcctttctgc aggtctccag aagcgggatc cgactgctct cagtgggtac tctcaatctg 1140
gagacgaacc agcaccagtt taggcatcct gacctctttg aggtgccaac cctcgaggca 1200
cccggggacg tggacccaaa ggggttgcac tctttcttag tgaatcagcc ctttgctgtc 1260
atcctcattg atcgcgagtc cggctgcgtc ctataccaag ccagaatcaa ggtgcctgag 1320
ccccccccca actccacact ctgccactaa 1350
<210> 4
<211> 1362
<212> DNA
<213> Taenia solium (Taenia saginata)
<400> 4
atgatttctc tacatatgag agacaaagga aaggcagctg catctgagcc tgagagaccc 60
aaatctactg tttcgaaaag cgtcaaagtc aatccggtcg actatctgaa gcacagggac 120
ttcaacaaca aatattgctt cggtgttcat gccattaatg acatcaccaa gaagtcgggt 180
cactcgccaa caacgatacg cttcctcctg acagtgttgg tgggctcgaa ggcggcacgg 240
ggcaccagtg cggaccagat cacacaggca ctcagcacca cgaactcacg tgatgtgggc 300
accgaatgca ccagcctcgt ggacagcgcg cttgatgagt gggctctgct ctctactgcg 360
ggtctaagcg atataccact cgatcgcatt gaggaagggc gcctctgtcg tttccagtct 420
gctatcttcg tccccatgga tgatctcacc ttcaaagccg agttccagtg gctgatcacc 480
aaccgcatgg gtgtcgtctg gactcagact tcgagaaatg atttcgccca tgctcagaag 540
tggttatcca aagtgtcaaa aggtttattc actcaccact ttccaaaaca atgtaccaac 600
tctcttgtac ttgccacagc attacaattt aaaggcaggt ggactcaacc tttggagctc 660
tacggtagtt ccaatggatc attcgaagtg tcaccccact cgaggataga tattcctatg 720
ctaaaaattt caacaaaggt tatttactac aaggatacca aaaaaggctt ccatttggtt 780
ggaatcccac ttaaggatac acggttcgca atcgcatttc tattaccctt ggtgccgcac 840
aagttcgtag aggtggagca tcgctttaca gagggtttga actacggtct cttcaacagc 900
tcgcgactgc atttctgcag catgcacgtg gtcattccta tgtttcgagt gcaaacggag 960
gtggacctct gtaaggcctt gcccttcctc ggcatgagca atccattcga ctctgatcgg 1020
gccgacttct ccggtatcag cgatgtagag aatctgcata tcaacagcgg caaggagtcg 1080
gcctttctgc aggtctccag aagcgggatc cgactgctct cagtgggtac tctcaatctg 1140
gagacgaacc agcaccagtt taggcatcct gacctctttg aggtgccaac tctcgaggca 1200
cccggggacg tggacccaaa ggaggtgcac tttttcttag tgaatcagcc ctttgctgtc 1260
atcctcattg atcgcgagtc cgactgcgtc ctataccaag ccagaatcaa ggtgcctgag 1320
cccctctcca actccacact gcctgccact aacttacaat aa 1362
<210> 5
<211> 1362
<212> DNA
<213> Asian tapeworm (Taenia asiatica)
<400> 5
atgatttctc tacatataag agacaaagga aaggcagctg catctgagcc tgagagaccc 60
aaatctactg tttcgaaaag cgtcaaagtc aatccggtcg actctctgaa gcacagggac 120
ttcaacaaca aatactgctt cggtgttcat gccattaatg acatcaccaa gaagtcgggt 180
cactcgccaa caacgatacg cttcctcctg acagtgttgg tgggctcgaa ggcggcacgg 240
ggcaccagtg cggaccagat cacacaggca ctcagcacca cgaactcacg tgatgtgggc 300
accgaatgca ccagcctcgt ggacagcgcg cttgatgagt gggctctgct ctctactgcg 360
ggtctaagcg atataccact cgatcgcatt gaggaagggc gcctctgtcg tttccagtct 420
gctatcttcg tccccatgga tgatctcacc ttcaaagccg agttccagtg gctgatcacc 480
aaccgcatgg gtgtcgtctg gactcagact tcgagaaatg atttcgccca tgctcagaag 540
tggttatcca aagtgtcaaa aggtttattc actcaccact ttccaaaaca atgtaccaac 600
tctcttgtac ttgccacagc attacaattt aaaggtaggt ggactcaacc tttggagctc 660
tacggtagtt ccaagggatc attcgaagtg tcaccccact cgaggataga tattcctatg 720
ctaaaaattt caacaaaggt tatttactac aaggatacca aaaaaggctt ccatttggtt 780
ggaatcccac ttaaggatac acggttcgca atcgcatttc tattaccctt ggtgccgcac 840
aagttcgtag aggtggagca tcgctttaca gagggtttga actacggtct cttcaacagc 900
tcgcgactgc atttctgcag catgcacgtg gtcattccta tgtttcgagt gcaaacggag 960
gtggacctct gtaaggcctt gcccttcctc ggcatgagca atccattcga ctctgatcgg 1020
gccgacttct ccggtatcag cgatgtagag aatctgcata tcaacagcgg caaggagtcg 1080
gcctttctgc aggtctccag aagcgggatc cgactgctct cagtgggtac tctcagtctg 1140
gagacgaacc agcaccagtt taggcatcct gacctctttg aggtgccaac cctcgaggca 1200
cccggggacg tggacccaaa ggaggtgcac tttttcttag tgaatcagcc ctttgctgtc 1260
atcctcattg atcgcgagtc cgactgcgtc ctataccaag ccagaatcaa ggtgcctgag 1320
cccctctcca actccacact gcctgccact aacttacaat aa 1362
Claims (5)
1. The method for identifying three human tapeworms for non-disease diagnosis and treatment is characterized by comprising the following steps:
(1) extracting three kinds of total RNA of the human tapeworm to be identified, and carrying out reverse transcription on the total RNA by using a universal primer as described below to synthesize cDNA of the human tapeworm to be identified;
the universal primer comprises an upstream primer shown as SEQ ID NO. 1 and a downstream primer shown as SEQ ID NO. 2; wherein the content of the first and second substances,
SEQ ID NO:1：5’-AGAGACAAAGGAAAGGCAGCTG-3’；
SEQ ID NO:2：5’-GGGGCTCAGGCACCTTGAT-3’；
(2) carrying out ep45 gene PCR amplification by taking the human tapeworm cDNA to be identified as a template; obtaining a PCR amplification product;
(3) carrying out agarose gel electrophoresis on the PCR amplification product, and then carrying out gel recovery; obtaining an ep45 glue recovery product;
(4) carrying out enzyme digestion treatment on the recovered product of the ep45 gel by using EcoRI and/or AarI;
(5) carrying out agarose gel electrophoresis on the enzyme digestion product, and identifying the human tapeworm according to the enzyme digestion map; the PCR amplification conditions of the ep45 gene are as follows: pre-denaturation at 95 ℃ for 5 min; denaturation at 94 ℃ for 1min, annealing at 60 ℃ for 1min, and extension at 72 ℃ for 90s for 30 cycles; after the end of the last cycle, the extension was carried out for 10min at 72 ℃.
2. The method for identifying three human tapeworms according to claim 1, wherein the reaction system for PCR amplification of the ep45 gene is as follows:
rTaq mixture：25.0μL；
SEQ ID NO:1：1.0μL；
SEQ ID NO:2：1.0μL；
and (3) identifying human tapeworm cDNA: 1.0 μ L;
DEPC H2O：22.0μL。
3. the method for identifying three human tapeworms of claim 1, wherein in step (3), the DNA is recovered and purified by gel electrophoresis.
4. The method for identifying three human tapeworms according to claim 1, wherein the total RNA of the human tapeworms to be identified is extracted from tapeworm segments or worm eggs.
5. The method for identifying three human tapeworms according to claim 1, wherein the enzyme digestion system of the ep45 gene is as follows:
the product recovered from the gel was 5. mu.L
EcoRI or AarI 1. mu.L
Buffer 2 μL
11.5 μ L deionized water.
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| CN201710647928.2A CN107326082B (en) | 2017-08-01 | 2017-08-01 | Universal primer of human cestode ep45 gene and identification method thereof |
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| CN107326082B true CN107326082B (en) | 2020-12-15 |
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| CN110082534A (en) * | 2019-03-20 | 2019-08-02 | 中国农业科学院兰州兽医研究所 | A kind of universal testing kit of sheep Babesia |
| CN110714087B (en) * | 2019-11-14 | 2021-10-26 | 中国农业科学院兰州兽医研究所 | Primer, kit and identification method for simultaneously and rapidly identifying 4 taenia solium larvae |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102424837A (en) * | 2011-12-09 | 2012-04-25 | 中国农业科学院兰州兽医研究所 | Kit for identifying and distinguishing types of human taeniasis |
| CN103952467A (en) * | 2014-02-18 | 2014-07-30 | 中国农业科学院兰州兽医研究所 | Special primers and identification method for rapidly identifying anoplocephalidae tapeworms parasitizing on Equine |
| CN104164512A (en) * | 2014-08-26 | 2014-11-26 | 中国农业科学院兰州兽医研究所 | PCR-RFLP detection kit for authenticating and differentiating infections of echinococcus multilocularis and echinococcosis shiquicus |
| CN106432482A (en) * | 2016-09-29 | 2017-02-22 | 中国农业科学院兰州兽医研究所 | Taenia solium TsEP45 recombinant protein and application thereof |
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| CN105924518B (en) * | 2016-04-20 | 2019-05-28 | 中国农业科学院兰州兽医研究所 | Taeniasis suis TsSerpin B6 recombinant protein and its application |
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Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102424837A (en) * | 2011-12-09 | 2012-04-25 | 中国农业科学院兰州兽医研究所 | Kit for identifying and distinguishing types of human taeniasis |
| CN103952467A (en) * | 2014-02-18 | 2014-07-30 | 中国农业科学院兰州兽医研究所 | Special primers and identification method for rapidly identifying anoplocephalidae tapeworms parasitizing on Equine |
| CN104164512A (en) * | 2014-08-26 | 2014-11-26 | 中国农业科学院兰州兽医研究所 | PCR-RFLP detection kit for authenticating and differentiating infections of echinococcus multilocularis and echinococcosis shiquicus |
| CN106432482A (en) * | 2016-09-29 | 2017-02-22 | 中国农业科学院兰州兽医研究所 | Taenia solium TsEP45 recombinant protein and application thereof |
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