CN112662784A - Atlantic salmon DNA bar code standard detection gene, primer and application thereof - Google Patents
Atlantic salmon DNA bar code standard detection gene, primer and application thereof Download PDFInfo
- Publication number
- CN112662784A CN112662784A CN202110106224.0A CN202110106224A CN112662784A CN 112662784 A CN112662784 A CN 112662784A CN 202110106224 A CN202110106224 A CN 202110106224A CN 112662784 A CN112662784 A CN 112662784A
- Authority
- CN
- China
- Prior art keywords
- atlantic salmon
- primer
- dna
- standard detection
- salmon
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Images
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention relates to a DNA barcode standard detection gene and primer for Atlantic salmon and application thereof, belonging to the field of molecular biology, wherein the sequence is SEQ ID NO:1 is shown. The invention also provides a pair of primers for detecting the Atlantic salmon DNA barcode standard detection gene, and application of the Atlantic salmon DNA barcode standard detection sequence in identification of Atlantic salmon. Amplifying the Atlantic salmon DNA by using the primer, carrying out nucleotide sequence determination on the obtained PCR amplification product, judging a species identification result according to a sequence comparison analysis result, and if the species identification result is identical to the species identification result shown as SEQ ID NO:1 is less than 0.02, the sample to be detected can be judged to be the Atlantic salmon. The method of the invention can distinguish Atlantic salmon from other salmon and trout.
Description
Technical Field
The invention belongs to the field of molecular biology, and particularly relates to a DNA barcode standard detection gene and primer for Atlantic salmon and application thereof.
Background
Atlantic Salmon (Salmon salar) belonging to Salmoniformes (Salmoniformes) Salmonidae (Salmonidae) Salmonella (Salmo) is a very typical migratory fish that is distributed mainly in the North Atlantic sea area and in northern Europe and the east coast of North America. They live most of the time in seawater, and their reproductive phase swims to rivers where they are born to lay eggs, and young fish also grow up in freshwater environment and then return to the ocean for fattening. Atlantic salmon contains rich unsaturated fatty acid and astaxanthin, has extremely high nutrition and health care values, becomes an internationally recognized high-grade aquatic product, and is an aquatic product fine variety with higher development and culture values.
Salmon circulating in the market of China is mainly imported Atlantic salmon, and has Norway, Chile, Denmark and the like in the production area, and the price is relatively expensive. In addition, other salmon and trout fishes such as rainbow trout and salmon are generally sold in domestic markets as salmon, and the selling price is relatively low. The meat quality of the salmon and the trout of different types is different, the breeding cost, the selling price and the nutritional value of the salmon and the trout are different, and the consumption preference of different countries is different, so that the phenomenon of confused selling causes considerable disputes. Most of salmon sold in the market are tissues such as muscles and bones, the species of salmon can be judged without complete morphology, and consumers cannot judge the species. Therefore, it is important to develop an efficient non-morphological method for identifying Atlantic salmon.
Disclosure of Invention
The technical problem to be solved by the invention is to provide a section of Atlantic salmon DNA barcode standard detection gene and application thereof in species identification. The DNA barcode standard detection sequence for the Atlantic salmon provided by the invention is beneficial to realizing the molecular identification of the Atlantic salmon, and can effectively shorten the identification time.
The Atlantic salmon DNA bar code standard detection gene is shown as SEQ ID NO:1 is shown.
The invention also provides primers for detecting the Atlantic salmon DNA barcode standard detection gene, which comprise a forward primer 5'-CTCTATTTAGTATTTGGTGCCTGA-3' and a reverse primer 5'-TAGACTTCTGGGTGGCCAAAGAACC-3'.
The application of the Atlantic salmon DNA barcode standard detection gene in identifying Atlantic salmon.
The specific identification method comprises the following steps:
1) separating and extracting DNA from a tissue sample to be detected;
2) performing PCR amplification by using the DNA obtained in the step 1) as a template and adopting a forward primer and a reverse primer;
3) separating the PCR amplification product obtained in the step 2) by agarose gel electrophoresis, judging the result according to the size of the amplification product, and if the result is identical to the result shown in SEQ ID NO:1 is less than 0.02, the sample to be detected can be judged to be the Atlantic salmon.
Further, the forward primer in step 2) is 5'-CTCTATTTAGTATTTGGTGCCTGA-3', and the reverse primer is 5'-TAGACTTCTGGGTGGCCAAAGAACC-3'.
Further, the conditions of the PCR amplification reaction in step 2) are: pre-denaturation at 95 ℃ for 3 min; denaturation at 95 ℃ for 15sec, annealing at 50 ℃ for 10sec, and extension at 72 ℃ for 40sec for 35 cycles; finally, extension was carried out at 72 ℃ for 7 min.
Compared with the prior art, the invention has the beneficial effects that:
the invention provides a DNA barcode standard detection sequence for identifying Atlantic salmon, which realizes rapid and accurate identification of Atlantic salmon and overcomes the following defects in the prior art: individuals with incomplete disabilities are essentially not identified, and the identification of complete individuals requires a professional who is well familiar with Atlantic salmon. Compared with the traditional morphological identification, the identification time is effectively shortened, and the identification method is accurate.
The method has accurate detection and can distinguish the Atlantic salmon from other fish species of the same genus.
In the identification method provided by the invention, the repeatability and the stability of a PCR reaction system and reaction conditions are high.
Drawings
FIG. 1 is an electrophoresis diagram of the PCR amplification product.
Detailed Description
The technical solution of the present invention is not limited to the following specific embodiments, but includes any combination of the specific embodiments.
Example 1
The DNA barcode standard detection gene of the Atlantic salmon of the embodiment is shown as SEQ ID NO:1 is shown.
Primers for detecting the Atlantic salmon DNA barcode standard detection gene include forward primer 5'-CTCTATTTAGTATTTGGTGCCTGA-3' and reverse primer 5'-TAGACTTCTGGGTGGCCAAAGAACC-3'.
The application of the Atlantic salmon DNA barcode standard detection gene in identifying Atlantic salmon. The specific identification method comprises the following steps:
firstly, separating and extracting DNA from a tissue sample to be detected;
secondly, performing PCR amplification by using the DNA obtained in the first step as a template and adopting a forward primer and a reverse primer; the conditions of the PCR amplification reaction are as follows: pre-denaturation at 95 ℃ for 3 min; denaturation at 95 ℃ for 15sec, annealing at 50 ℃ for 10sec, and extension at 72 ℃ for 40sec for 35 cycles; finally, extension was carried out at 72 ℃ for 7 min.
And thirdly, separating the PCR amplification product obtained in the second step by agarose gel electrophoresis, judging the result according to the size of the amplification product, and if the result is identical to the result shown in SEQ ID NO:1 is less than 0.02, the sample to be detected can be judged to be the Atlantic salmon.
Example 2:
1) extracting DNA of a detection sample:
selecting 10 detection samples, wherein S1, S2 and S3 are Atlantic salmon (Salmo salar) from Chile; s4, S5, S6 are atlantic salmon in norway; s7 and S8 are Norway rainbow trout (Oncorhynchus mykiss); s9 is salmon (Oncorhynchus keta) of America; s10 is Chile' S silver salmon (Oncorhynchus kisutch). The above four fishes belong to Salmoniformes (Salmoniformes) Salmonidae (Salmonoidae).
The DNA was extracted according to the instruction of the DNA extraction kit (centrifugal column type) (Tiangen Biochemical technology Co., Ltd.) for marine organism tissue. Cutting tissue material not more than 30mg, placing into a centrifuge tube containing 200 μ lGA buffer solution, and vortex-oscillating for 15 sec; adding 20 μ l of ProteinaseK (20mg/ml) solution, mixing by vortexing, and centrifuging briefly to remove water droplets on the inner wall of the tube cover; standing at 56 deg.C until the tissue is completely dissolved, and centrifuging briefly to remove water droplets on the inner wall of the tube cover; adding 200 μ l buffer solution GB, fully reversing and mixing, standing at 70 deg.C for 10min, cleaning the solution, and centrifuging briefly to remove water droplets on the inner wall of the tube cover; adding 200 μ l of anhydrous ethanol, mixing thoroughly, wherein flocculent precipitate may appear, and centrifuging briefly to remove water drops on the inner wall of the tube cover; adding the solution and flocculent precipitate obtained in the previous step into an adsorption column CB3 (the adsorption column is placed into a collecting pipe), centrifuging at 12,000rpm (-13,400 Xg) for 30sec, pouring the waste liquid, and placing the adsorption column CB3 back into the collecting pipe; adding 500 μ l of buffer GD into adsorption column CB3, centrifuging at 12,000rpm (-13,400 Xg) for 30sec, pouring off waste liquid, and placing adsorption column CB3 into a collection tube; adding 600 μ l of rinsing liquid PW into adsorption column CB3, centrifuging at 12,000rpm (-13,400 Xg) for 30sec, pouring off waste liquid, and placing adsorption column CB3 into a collecting tube; repeating the previous step; the adsorption column CB3 is put back into the collecting tube, centrifuged for 2min at 12,000rpm (13,400 Xg), and the waste liquid is poured out; placing the adsorption column CB3 at room temperature for a plurality of minutes to thoroughly dry the residual rinsing liquid in the adsorption material; transferring the adsorption column CB3 into a clean centrifugal tube, hanging and dripping 50-200 mu l of elution buffer TE into the middle part of the adsorption film, placing the centrifugal tube for 2-5min at room temperature, centrifuging the centrifugal tube for 2min at 12,000rpm (13,400 Xg), collecting the solution into the centrifugal tube, and placing the centrifugal tube for storage at 40 ℃ for later use, thus completing the DNA extraction of the sample to be detected;
2) and (3) PCR amplification:
the PCR reaction system is 25 mul and consists of 2 x Rapid Taq Master Mix 12.5 mul, forward and reverse primers of 10.0mmol/L each 1 mul, DNA template 2 mul and deionized water 8.5 mul; the reaction procedure for PCR was: pre-denaturation at 95 ℃ for 3 min; denaturation at 95 ℃ for 15sec, annealing at 50 ℃ for 10sec, and extension at 72 ℃ for 40sec for 35 cycles; finally, extension was carried out at 72 ℃ for 7 min.
The primers include a forward primer 5'-CTCTATTTAGTATTTGGTGCCTGA-3' and a reverse primer 5'-TAGACTTCTGGGTGGCCAAAGAACC-3'.
3) Sequencing of test samples
The PCR amplification product is a 680bp band, and the electrophoretogram is shown in FIG. 1. The PCR product is sent to a biological company for sequencing, so as to obtain ten sequencing sample sequences S1, S2, S3, S4, S5, S6, S7, S8, S9 and S10, wherein the sequences are SEQ ID NO: 4-13.
5) Alignment of Gene sequences
Pairwise alignment of the Atlantic salmon DNA barcode standard sequence (shown as SEQ ID NO:1 in the sequence table) and ten sequencing sample sequences (S1, S2, S3, S4, S5, S6, S7, S8, S9 and S10) by using biological software BioEdit shows that the genetic distances of the Atlantic salmon standard gene sequence and the six Atlantic salmon sequencing sequences from different sources S1, S2, S3, S4, S5 and S6 after alignment are respectively 0.006 and are less than 0.02. The genetic distances to four sample sequencing sequences S7, S8, S9 and S10 of rainbow trout, salmon and silver salmon are respectively 0.151, 0.163 and 0.149, and are all more than 0.02. The six sequencing sequences S1, S2, S3, S4, S5 and S6 can be judged to be Atlantic salmon, and the six tissues to be tested can also be identified as the tissues of the Atlantic salmon. The comparison results are shown in Table 1, LW in Table 1 is Atlantic salmon DNA barcode standard sequence (SEQ ID NO: 1); SEQ ID NO: 4-13 are the nucleotide sequences of 10 detection samples respectively.
TABLE 1
| S1 | S2 | S3 | S4 | S5 | S6 | S7 | S8 | S9 | S10 | AS | |
| S1 | |||||||||||
| S2 | 0.006 | ||||||||||
| S3 | 0.006 | 0.000 | |||||||||
| S4 | 0.006 | 0.000 | 0.000 | ||||||||
| S5 | 0.006 | 0.000 | 0.000 | 0.000 | |||||||
| S6 | 0.006 | 0.000 | 0.000 | 0.000 | 0.000 | ||||||
| S7 | 0.151 | 0.149 | 0.149 | 0.149 | 0.149 | 0.149 | |||||
| S8 | 0.151 | 0.149 | 0.149 | 0.149 | 0.149 | 0.149 | 0.003 | ||||
| S9 | 0.163 | 0.163 | 0.163 | 0.163 | 0.163 | 0.163 | 0.107 | 0.107 | |||
| S10 | 0.149 | 0.147 | 0.147 | 0.147 | 0.147 | 0.147 | 0.070 | 0.070 | 0.105 | ||
| AS | 0.003 | 0.006 | 0.006 | 0.006 | 0.006 | 0.006 | 0.153 | 0.153 | 0.167 | 0.151 |
Sequence listing
<110> research institute for aquatic products in yellow sea of China institute for aquatic science
<120> Atlantic salmon DNA barcode standard detection gene, primer and application thereof
<160> 13
<170> SIPOSequenceListing 1.0
<210> 1
<211> 680
<212> DNA
<213> Atlantic salmon (Salmo salar)
<400> 1
ctctatttag tatttggtgc ctgagccgga atagtcggca ccgccctaag tctcttgatt 60
cgagcagaac tcagccagcc tggcgccctt ctgggagatg accaaattta taacgtaatt 120
gttacagccc atgccttcgt cataattttc tttatagtca taccgattat gatcggcggc 180
tttggaaact gattaattcc tcttataatc ggggcccccg acatagcatt cccccgaatg 240
aataacataa gtttttgact tctccctccc tcctttcttc tcctcctggc ctcatctgga 300
gttgaagccg gcgctggcac cggatgaaca gtctaccccc ctctagcagg taatcttgcc 360
cacgcaggag cttccgttga cttaactatt ttttccctcc atttggctgg tatttcttca 420
attcttgggg ccattaattt tatcacaacc attattaata taaaaccccc agctatctct 480
cagtatcaaa ccccactttt tgtttgagct gtattagtca ctgccgtcct tttattactc 540
tccctccctg ttctagcagc aggcattacc atactactta cagaccgaaa tctaaatacc 600
actttctttg acccggcagg cggaggggac ccaatcttgt accaacatct cttttggttc 660
tttggccacc cagaagtcta 680
<210> 2
<211> 24
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 2
ctctatttag tatttggtgc ctga 24
<210> 3
<211> 25
<212> DNA
<213> Artificial Sequence (Artificial Sequence)
<400> 3
tagacttctg ggtggccaaa gaacc 25
<210> 4
<211> 680
<212> DNA
<213> Atlantic salmon (Salmo salar)
<400> 4
ctctatttag tatttggtgc ctgagccgga atagtcggca ccgccctaag tctcttgatt 60
cgagcagaac tcagccagcc tggcgccctt ctgggagatg accaaattta taacgtaatt 120
gttacagccc atgccttcgt cataattttc tttatagtca taccgattat gatcggcggc 180
tttggaaact gattaattcc tcttataatc ggggcccccg acatagcatt cccccgaatg 240
aataacataa gtttttgact tctccctccc tcctttcttc tcctcctggc ctcatctgga 300
gttgaagccg gcgctggcac cggatgaaca gtctaccccc ctctagcagg taatcttgcc 360
cacgcaggag cttccgttga cttaactatt ttttccctcc atttggctgg tatttcttca 420
attctggggg ccattaattt tatcacaacc attattaata taaaaccccc agctatctct 480
cagtatcaaa ccccactttt tgtttgagct gtattagtca ctgccgtcct tttattactc 540
tccctccctg ttctagcagc aggcattacc atactactta cagaccgaaa tctaaatacc 600
actttctttg acccggcagg cggaggggac ccaatcttgt accaacatct cttttggttc 660
tttggccacc cagaagtcta 680
<210> 5
<211> 680
<212> DNA
<213> Atlantic salmon (Salmo salar)
<400> 5
gacattggca ccctctattt agtatttggt gcctgagccg gaatagtcgg caccgcccta 60
agtctcttga ttcgagcaga actcagccag cctggcgccc ttctgggaga tgaccaaatt 120
tataacgtaa ttgttacagc ccatgccttc gtcataattt tctttatagt cataccgatt 180
atgatcggcg gctttggaaa ctgattaatt cctcttataa tcggggcccc cgacatagca 240
ttcccccgaa tgaataacat aagtttttga cttctccctc cctcctttct tctcctcctg 300
gcctcatctg gagttgaagc cggcgctggc accggatgaa cagtctaccc ccctctagca 360
ggtaatcttg cccacgcagg agcttccgtt gacttaacta ttttttccct ccatttggct 420
ggtatttctt caattcttgg ggccattaat tttattacaa ccattattaa tataaaaccc 480
ccagctatct ctcagtatca aaccccactt tttgtttgag ctgtattagt cactgccgtc 540
cttttgttac tctccctccc tgttctagca gcaggcatta ccatactact tacagaccga 600
aatctaaata ccactttctt tgacccggca ggcggaggag acccaatctt cttttggttc 660
ctcttttgat tctttggcca 680
<210> 6
<211> 680
<212> DNA
<213> Atlantic salmon (Salmo salar)
<400> 6
gacattggca ccctctattt agtatttggt gcctgagccg gaatagtcgg caccgcccta 60
agtctcttga ttcgagcaga actcagccag cctggcgccc ttctgggaga tgaccaaatt 120
tataacgtaa ttgttacagc ccatgccttc gtcataattt tctttatagt cataccgatt 180
atgatcggcg gctttggaaa ctgattaatt cctcttataa tcggggcccc cgacatagca 240
ttcccccgaa tgaataacat aagtttttga cttctccctc cctcctttct tctcctcctg 300
gcctcatctg gagttgaagc cggcgctggc accggatgaa cagtctaccc ccctctagca 360
ggtaatcttg cccacgcagg agcttccgtt gacttaacta ttttttccct ccatttggct 420
ggtatttctt caattcttgg ggccattaat tttattacaa ccattattaa tataaaaccc 480
ccagctatct ctcagtatca aaccccactt tttgtttgag ctgtattagt cactgccgtc 540
cttttgttac tctccctccc tgttctagca gcaggcatta ccatactact tacagaccga 600
aatctaaata ccactttctt tgacccggca ggcggaggag acccaatctt cttttggttc 660
ctcttttgat tctttggcca 680
<210> 7
<211> 680
<212> DNA
<213> Atlantic salmon (Salmo salar)
<400> 7
gacattggca ccctctattt agtatttggt gcctgagccg gaatagtcgg caccgcccta 60
agtctcttga ttcgagcaga actcagccag cctggcgccc ttctgggaga tgaccaaatt 120
tataacgtaa ttgttacagc ccatgccttc gtcataattt tctttatagt cataccgatt 180
atgatcggcg gctttggaaa ctgattaatt cctcttataa tcggggcccc cgacatagca 240
ttcccccgaa tgaataacat aagtttttga cttctccctc cctcctttct tctcctcctg 300
gcctcatctg gagttgaagc cggcgctggc accggatgaa cagtctaccc ccctctagca 360
ggtaatcttg cccacgcagg agcttccgtt gacttaacta ttttttccct ccatttggct 420
ggtatttctt caattcttgg ggccattaat tttattacaa ccattattaa tataaaaccc 480
ccagctatct ctcagtatca aaccccactt tttgtttgag ctgtattagt cactgccgtc 540
cttttgttac tctccctccc tgttctagca gcaggcatta ccatactact tacagaccga 600
aatctaaata ccactttctt tgacccggca ggcggaggag acccaatctt gtattggttc 660
ctcttttgat tctttggcca 680
<210> 8
<211> 670
<212> DNA
<213> Atlantic salmon (Salmo salar)
<400> 8
ctctatttag tatttggtgc ctgagccgga atagtcggca ccgccctaag tctcttgatt 60
cgagcagaac tcagccagcc tggcgccctt ctgggagatg accaaattta taacgtaatt 120
gttacagccc atgccttcgt cataattttc tttatagtca taccgattat gatcggcggc 180
tttggaaact gattaattcc tcttataatc ggggcccccg acatagcatt cccccgaatg 240
aataacataa gtttttgact tctccctccc tcctttcttc tcctcctggc ctcatctgga 300
gttgaagccg gcgctggcac cggatgaaca gtctaccccc ctctagcagg taatcttgcc 360
cacgcaggag cttccgttga cttaactatt ttttccctcc atttggctgg tatttcttca 420
attcttgggg ccattaattt tattacaacc attattaata taaaaccccc agctatctct 480
cagtatcaaa ccccactttt tgtttgagct gtattagtca ctgccgtcct tttgttactc 540
tccctccctg ttctagcagc aggcattacc atactactta cagaccgaaa tctaaatacc 600
actttctttg acccggcagg cggaggagac ccaatcttgt accaacatct cttttggttc 660
tttggccacc 670
<210> 9
<211> 670
<212> DNA
<213> Atlantic salmon (Salmo salar)
<400> 9
ctctatttag tatttggtgc ctgagccgga atagtcggca ccgccctaag tctcttgatt 60
cgagcagaac tcagccagcc tggcgccctt ctgggagatg accaaattta taacgtaatt 120
gttacagccc atgccttcgt cataattttc tttatagtca taccgattat gatcggcggc 180
tttggaaact gattaattcc tcttataatc ggggcccccg acatagcatt cccccgaatg 240
aataacataa gtttttgact tctccctccc tcctttcttc tcctcctggc ctcatctgga 300
gttgaagccg gcgctggcac cggatgaaca gtctaccccc ctctagcagg taatcttgcc 360
cacgcaggag cttccgttga cttaactatt ttttccctcc atttggctgg tatttcttca 420
attcttgggg ccattaattt tattacaacc attattaata taaaaccccc agctatctct 480
cagtatcaaa ccccactttt tgtttgagct gtattagtca ctgccgtcct tttgttactc 540
tccctccctg ttctagcagc aggcattacc atactactta cagaccgaaa tctaaatacc 600
actttctttg acccggcagg cggaggagac ccaatcttgt accaacatct cttttggttc 660
tttggccacc 670
<210> 10
<211> 665
<212> DNA
<213> Norway rainbow trout (Oncorhynchus mykiss)
<400> 10
ctctatttag tatttggtgc ctgagccggg atagtaggca ccgccctgag tctactgatt 60
cgggcggaac taagccagcc gggcgctctt ctgggggatg accaaatcta taacgtgatc 120
gtcacagccc atgccttcgt tatgattttc tttatagtca tgccaattat aatcgggggc 180
tttggaaact gattaattcc cttaataatc ggagcccctg atatggcatt ccctcgaata 240
aataacataa gcttctgact ccttcctcca tcctttctcc tcctcctgtc ttcatcagga 300
gttgaagccg gcgcgggtac tggatgaaca gtataccccc ctctagccgg caacctcgcc 360
cacgcaggag cctctgttga tttaactatc ttctcccttc atttagctgg aatctcctca 420
attttaggag ccattaattt tattacgacc attattaaca taaaacctcc agccatctct 480
cagtaccaaa cccccctttt cgtttgagcc gtgctagtta ctgctgtcct tctattactt 540
tccctccccg tcctggcagc aggcattact atgttactta cagaccgaaa tctaaacacc 600
actttctttg acccggcagg cgggggagat ccaattttat accaacacct cttttggttc 660
tttgg 665
<210> 11
<211> 665
<212> DNA
<213> Norway rainbow trout (Oncorhynchus mykiss)
<400> 11
ctctatttag tatttggtgc ctgagccggg atagtaggca ccgccctgag tctactgatt 60
cgggcggaac taagccagcc gggcgctctt ctgggggatg accaaatcta taacgtgatc 120
gtcacagccc atgccttcgt tatgattttc tttatagtca tgccaattat aatcgggggc 180
tttggaaact gattaattcc cctaataatc ggagcccctg atatggcatt tcctcgaata 240
aataacataa gcttctgact ccttcctcca tcctttctcc tcctcctgtc ttcatcagga 300
gttgaagccg gcgcgggtac tggatgaaca gtataccccc ctctagccgg caacctcgcc 360
cacgcaggag cctctgttga tttaactatc ttctcccttc atttagctgg aatctcctca 420
attttaggag ccattaattt tattacgacc attattaaca taaaacctcc agccatctct 480
cagtaccaaa cccccctttt cgtttgagcc gtgctagtta ctgctgtcct tctattactt 540
tccctccccg tcctggcagc aggcattact atgttactta cagaccgaaa tctaaacacc 600
actttctttg acccggcagg cgggggagat ccaattttat accaacacct cttttgattc 660
tttgg 665
<210> 12
<211> 665
<212> DNA
<213> salmon in the United states (Oncorhynchus keta)
<400> 12
ctctatttag tatttggtgc ctgagccggg atagtaggca ccgccctgag cctactaatt 60
cgggcagaac taagccagcc aggcgctctt ctaggggatg accagatcta caacgtaatc 120
gttacagccc atgccttcgt tataattttc tttatagtca taccaattat aatcggaggc 180
tttggaaact gattaatccc cctaatgatc ggggcaccag atatagcatt cccacgaata 240
aataacataa gcttctgact cctacctccg tccttcctcc tcctcctttc ttcatctgga 300
gttgaagccg gcgctggtac cgggtggaca gtttaccccc ctctagccgg aaaccttgcc 360
cacgcaggag catctgtcga cttaaccatc ttctccctcc atttagctgg aatctcctca 420
attttggggg ccattaattt tattacgacc attatcaaca taaaaccccc agctatttct 480
cagtaccaaa ccccgctttt tgtctgagct gtactaatca ctgctgtact tctactatta 540
tcactccccg ttctggcagc aggtattact atgttgctca cagatgggaa atttaaacac 600
cactttcttt gacccagcgg gcggcggaga tcccaatttt ataccaacac ctcttttgat 660
tcttt 665
<210> 13
<211> 665
<212> DNA
<213> silver salmon of Chile (Oncorhynchus kisutch)
<400> 13
ctctatttag tatttggtgc ctgagccggg atagtaggca ccgccctaag tctactgatt 60
cgagcagaac tgagccagcc gggcgctctt ctaggggatg atcagattta caacgtaatc 120
gtcacagccc atgccttcgt tatgattttc tttatagtca tgccgattat gatcggaggc 180
tttggaaact gattaattcc cctaatgatc ggagcccctg atatggcatt ccctcgaata 240
aataacataa gcttctgact ccttccgcca tcctttctcc tcctcctatc ttcctctgga 300
gttgaagccg gggctggcac cgggtgaaca gtttatcccc ctctggccgg caacctcgcc 360
cacgcaggag cctcagttga tctgactatc ttctcccttc atttagccgg gatctcctca 420
attttaggag ccattaattt tattacgacc attattaaca taaagccccc agctatctct 480
cagtaccaaa ccccactttt tgtttgagct gtgctagtca ctgctgttct tctactactc 540
tctctccccg ttctggcagc aggcattact atgttactta cagaccgaaa tctaaacacc 600
actttctttg acccggcagg cgggggagat ccaattttat accaacacct cttttgattc 660
tttgg 665
Claims (5)
1. The Atlantic salmon DNA bar code standard detection gene is characterized in that the sequence of the gene is SEQ ID NO:1 is shown.
2. The primer for detecting the Atlantic salmon DNA barcode standard detection gene of claim 1, wherein the primer comprises a forward primer 5'-CTCTATTTAGTATTTGGTGCCTGA-3' and a reverse primer 5'-TAGACTTCTGGGTGGCCAAAGAACC-3'.
3. The use of the Atlantic salmon DNA barcode standard detection gene of claim 1 for identifying Atlantic salmon, which is characterized by comprising the following steps:
1) separating and extracting DNA from a tissue sample to be detected;
2) using the DNA obtained in the step 1) as a template, adopting a forward primer and a reverse primer to carry out PCR amplification,
3) separating the PCR amplification product obtained in the step 2) by agarose gel electrophoresis, judging the result according to the size of the amplification product, and if the result is identical to the result shown in SEQ ID NO:1 is less than 0.02, the sample to be detected can be judged to be the Atlantic salmon.
4. The use according to claim 3, wherein the forward primer of step 2) is 5'-CTCTATTTAGTATTTGGTGCCTGA-3' and the reverse primer 5'-TAGACTTCTGGGTGGCCAAAGAACC-3'.
5. The use according to claim 3, wherein the PCR amplification reaction of step 2) is performed under the following conditions: pre-denaturation at 95 ℃ for 3 min; denaturation at 95 ℃ for 15sec, annealing at 50 ℃ for 10sec, and extension at 72 ℃ for 40sec for 35 cycles; finally, extension was carried out at 72 ℃ for 7 min.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110106224.0A CN112662784A (en) | 2021-01-26 | 2021-01-26 | Atlantic salmon DNA bar code standard detection gene, primer and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110106224.0A CN112662784A (en) | 2021-01-26 | 2021-01-26 | Atlantic salmon DNA bar code standard detection gene, primer and application thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN112662784A true CN112662784A (en) | 2021-04-16 |
Family
ID=75415912
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202110106224.0A Pending CN112662784A (en) | 2021-01-26 | 2021-01-26 | Atlantic salmon DNA bar code standard detection gene, primer and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN112662784A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2022263600A1 (en) * | 2021-06-17 | 2022-12-22 | Salmotrace As | Method of tagging fish and other animals |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103122386A (en) * | 2013-01-29 | 2013-05-29 | 中国水产科学研究院黄海水产研究所 | Molecular identification method of atlantic salmon |
-
2021
- 2021-01-26 CN CN202110106224.0A patent/CN112662784A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103122386A (en) * | 2013-01-29 | 2013-05-29 | 中国水产科学研究院黄海水产研究所 | Molecular identification method of atlantic salmon |
Non-Patent Citations (3)
| Title |
|---|
| 孟玮等: "基于线粒体COI基因序列的亚东鲑DNA条形码研究", 《水产学杂志》 * |
| 李新光: "基于DNA条形码的鱼片（肉）真伪鉴别技术研究", 《中国优秀博硕士学位论文全文数据库(硕士)工程科技Ⅰ辑》 * |
| 殷军: "《全国高等医药院校药学类实验教材 生药学实验 第3版》", 31 August 2019, 中国医药科技出版社 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2022263600A1 (en) * | 2021-06-17 | 2022-12-22 | Salmotrace As | Method of tagging fish and other animals |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN113502333B (en) | Molecular marker C42257 for rapidly identifying genetic sex of penaeus japonicus and application thereof | |
| CN106520939B (en) | A kind of method and its application of madai Germplasm Identification | |
| CN112226518B (en) | Molecular marker C69483 for rapidly identifying genetic sex of red swamp crayfish and application thereof | |
| CN108179200B (en) | Microsatellite marker linked with procambarus clarkii high fertility character and application thereof | |
| CN111254202A (en) | Primer group, kit and detection method for detecting Atlantic salmon | |
| CN110760595B (en) | Identification primer and method suitable for tridacna species common in trioyster and south China sea and hybridized species of tridacna species | |
| CN112662784A (en) | Atlantic salmon DNA bar code standard detection gene, primer and application thereof | |
| CN105176989A (en) | Primers and method for identifying takifugu obscurus fry and takifugu xanthopterus fry | |
| CN111057771B (en) | SNP molecular marker for distinguishing 'Zhongyang No. 1' from common fugu obscurus and application thereof | |
| CN103555847A (en) | Method for paternity identification of tilapia mossambica | |
| CN111154852B (en) | Specific DNA fragment for sex identification of spiny eels and application thereof | |
| CN112226497A (en) | Molecular marker C98284 for rapidly identifying genetic sex of red swamp crayfish and application thereof | |
| CN113502334B (en) | Molecular marker C27449 for rapidly identifying genetic sex of Penaeus japonicus and application thereof | |
| CN104611440B (en) | A kind of method that parentage identification and paternity identification are carried out using multiple haliotis discus hannai Ino fluorescence labeling microsatellite | |
| CN110042169B (en) | Molecular marker primer, kit and identification method for group specificity of Fennel fish in Heilongjiang | |
| KR102083306B1 (en) | Genetic markers and methods for identifying species belong to Myxinidae | |
| CN108841930B (en) | Paramisgurnus dabryanus microsatellite family identification method and application thereof | |
| CN110607377A (en) | Primer, probe and method for identifying Antarctic canine fish based on 16S rRNA gene | |
| CN110592239A (en) | Primer, probe and method for identifying Antarctic canis based on Cyt b gene | |
| CN113621709B (en) | Microsatellite marked primer for yellow river carp and method for evaluating release effect of yellow river carp | |
| CN108660220A (en) | One group of sturgeon SNP marker, sturgeon production of hybrid seeds appraisal procedure and genetic evaluation method | |
| CN110564870A (en) | Primer, probe and method for identifying naked-capped fish based on Cytb gene | |
| CN113373244B (en) | Method for rapidly detecting genetic sex of channel catfish based on SNP locus specific primer extension reaction | |
| KR102367357B1 (en) | PCR primer sets and adapters for amplifying and sequencing mitochondrial ribosomal RNA gene used for DNA barcoding and metabarcoding of fishes | |
| CN114350815B (en) | Molecular marking method for identifying red sea bream and red sea bream filial generation by ISSR primer |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20210416 |
|
| RJ01 | Rejection of invention patent application after publication |