CN109055571A - The specific primer of yellowfin spine porgy microsatellite marker and application - Google Patents
The specific primer of yellowfin spine porgy microsatellite marker and application Download PDFInfo
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Abstract
The invention discloses a kind of specific primer of yellowfin spine porgy microsatellite marker, the specific primer includes 8 pairs of primers, respectively AL15, AL20, AL51, AL01, AL37, AL18, AL14 and AL49, and the primer polymorphism is high, and PCR product is reliable and stable；The invention also discloses a kind of yellowfin spine porgy microsatellite paternity test methods, this method establishes the paternity test molecular method of yellowfin spine porgy for the first time, this method identification is accurate, cheap, and the present invention further discloses application of the specific primer of above-mentioned yellowfin spine porgy microsatellite marker in yellowfin spine porgy paternity test.
Description
Technical field
The invention belongs to micro-satellite labeling technique fields, and in particular to a kind of specificity of yellowfin spine porgy microsatellite marker is drawn
Object and application.
Background technique
Yellowfin spine porgy (Acanthopagrus latus) is under the jurisdiction of net-rope Pisces, Perciformes Perciformes, porgy section
Sparidae, spine porgy belong to Acanthopagrus, be distributed widely in Red sea, Arabian Sea, the Indian Ocean, Western Pacific it is littoral and in
State Taiwan, Fujian, Guangdong, ALONG GUANGXI COAST.
Yellowfin spine porgy moves in immediate offshore area and firth, and omnivorousness is a kind of important economic fish.Yellowfin spine porgy is supported
It grows the period to grow partially, needs a year and a half to can be only achieved 5 liang or so of listing specification to 2 years, strongly limit the cultivation of the fisherman kind
Enthusiasm.In recent years, since environmental pollution, overfishing etc. lead to yellowfin spine porgy Natural Population slump of disastrous proportions, germplasm is seriously moved back
Change etc.；The reasons such as inbreeding, small dimension parent's artificial breeding of yellowfin spine porgy cultured population simultaneously cause yellowfin spine porgy to cultivate
Germplasm is seriously degenerated, and premunition weakens, culture performance decline etc..Problem above seriously constrains continuing for yellowfin spine porgy aquaculture
It develops in a healthy way.It is therefore extremely urgent to the protection and fine-variety breeding of the yellowfin spine porgy progress germ plasm resource of Different population,
And one of fine-variety breeding important method is exactly to carry out family selective breeding.Family selective breeding can obtain the kind with merit,
Simultaneously can accurately understand pedigree information, effectively instruct parental line selection, this can constantly improve inhereditary feature and
Breeding time is shortened in inbreeding, improves the yield of yellowfin spine porgy.
And in genetics-breeding in fish research, the source of seed is more mixed and disorderly, is difficult to area only with physics and morphological markers
Divide the seed of separate sources, different qualities.Generally, aquatic livestock is to carry out electronic marker with the identification method of pond mixed breeding family
Distinguish, but carrying out large-scale groups breeding or when family selective breeding, electronic marker it is expensive, therefore have certain limitation
Property.And DNA molecular marker technology has the characteristics that hereditary information is abundant, detection means is fast and convenient, reproducible, therefore is dividing
Sub- marker-assisted breeding and pedigree identification etc. are used widely.
With the rapid development of sequencing technologies, a large amount of DNA molecular marker such as microsatellite marker (Simple sequence
Repeat, SSR) and single nucleotide polymorphism (Single-nucleotide polymorphisms, SNPs) to be applied to aquatic products dynamic
Object, such as Parargyrops edita Tanaka (Parargyrops edita), carp (Cyprinus carpio), Larimichthys crocea (Larimichthy
Crocea) etc..These SSR markers and SNPs can be used to affiliation between clear individual, establish correct genealogical relationship, so both
The screening and popularization for being conducive to excellent breed variety are also beneficial to the seed market of specification China yellowfin spine porgy.Currently, existing one
A little scholars develop a large amount of yellowfin spine porgy SSR marker, but SSR marker is applied to the report of yellowfin spine porgy Parentage determination research
Road is very few, this is unfavorable for the process of yellowfin spine porgy fine-variety breeding.
Summary of the invention
The purpose of the present invention is to provide a kind of specific primer of yellowfin spine porgy microsatellite marker, the primer polymorphisms
Height, PCR product are reliable and stable.
The object of the invention is also to provide a kind of yellowfin spine porgy microsatellite paternity test method, this method identification is accurate,
It is cheap.
Final object of the present invention is the specific primer for providing above-mentioned yellowfin spine porgy microsatellite marker in Huang
Application in fin spine porgy paternity test.
Above-mentioned first purpose of the invention is achieved through the following technical solutions: a kind of yellowfin spine porgy microsatellite mark
The specific primer of note, the specific primer include 8 pairs of primers, respectively AL15, AL20, AL51, AL01, AL37, AL18,
AL14 and AL49, in which:
The nucleotide sequence of primer pair AL15 is as shown in SEQ ID NO:1 and SEQ ID NO:2；
The nucleotide sequence of primer pair AL20 is as shown in SEQ ID NO:3 and SEQ ID NO:4；
The nucleotide sequence of primer pair AL51 is as shown in SEQ ID NO:5 and SEQ ID NO:6；
The nucleotide sequence of primer pair AL01 is as shown in SEQ ID NO:7 and SEQ ID NO:8；
The nucleotide sequence of primer pair AL37 is as shown in SEQ ID NO:9 and SEQ ID NO:10；
The nucleotide sequence of primer pair AL18 is as shown in SEQ ID NO:11 and SEQ ID NO:12；
The nucleotide sequence of primer pair AL14 is as shown in SEQ ID NO:13 and SEQ ID NO:14；
The nucleotide sequence of primer pair AL49 is as shown in SEQ ID NO:15 and SEQ ID NO:16.
Above-mentioned second purpose of the invention is achieved through the following technical solutions: a kind of yellowfin spine porgy microsatellite parent
Sub- identification method, comprising the following steps:
(1) yellowfin spine porgy DNA is extracted: being acquired yellowfin spine porgy parental animal fin ray and the full fish of filial generation to be identified, is extracted base
Because of a group DNA；
(2) polymorphic micro-satellite markers screening and primer synthesis: screen and synthesize 8 pairs of above-mentioned primers, 8 pairs of primers
Be divided into three groups, wherein group one include AL15, AL20 and AL51, group two include AL01, AL37 and AL18, group three include AL14 and
AL49, group one and group two hold fluorophors different with tri- kinds of FAM, HEX and TAMRA respectively to be repaired the 5 ' of forward primer
Decorations, group three are modified respectively with the two different fluorophors of FAM and HEX at 5 ' ends of forward primer, are analyzed for PCR；
(3) step of three groups of primer pairs in step (2) microsatellite locus Genotyping: is utilized respectively using Fluorescence PCR
Suddenly the genomic DNA in (1) carries out PCR amplification, then mixes amplified production；
(4) Genotyping is carried out to mixed amplified production, using genotypic results to parent genotype and filial generation
Genotype is analyzed, and determines the Parent of offspring individual.
In above-mentioned yellowfin spine porgy microsatellite paternity test method:
In step (3) when Fluorescence PCR, every group of reaction system total volume is 10 μ L, comprising: distilled water 7.6 μ L, 10 ×
1 μ L of PCR (TaKaRa) buffer (contains Mg2+), concentration is 0.15 μ L, the rTaqDNA polymerase of dNTPs of 10mmol/L
(TaKaRa) 0.15 μ L, each 0.3 μ L of forward and reverse primer, concentration are the 0.5 μ L of genomic DNA of 100ng/ μ L.
In step (3) when Fluorescence PCR, PCR reaction condition are as follows: 95 DEG C of initial denaturation 4min；95 DEG C of denaturation 30s, annealing
Temperature 60 C 30s, 72 DEG C of extension 40min, 30 recycle；72 DEG C of extension 10min；4 DEG C of preservations.
In step (4) Genotyping is preferably carried out to mixed amplified production on ABI3730XL Genetic Analyser, used
GS-500LIZ reads the genotype of individual with 3.2 software of GeneMapper V as internal reference.
Preferably parent genotype and progeny genotypes are analyzed using 3.0 software of CERVUS in step (4), determined
The Parent of offspring individual.
Using software CERVUS3.0 can calculate parent and filial generation on each microsatellite seat gene frequency, heterozygosity,
It is expected that the letter such as heterozygosity, polymorphism information content, average probability of exclusion, Hardy-Weinberg balance and amorph frequency
Breath, therefore, it is determined that the Parent of offspring individual.
Further, a kind of yellowfin spine porgy microsatellite paternity test method provided by the invention, includes the following steps:
(1) yellowfin spine porgy DNA is extracted
The good yellowfin spine porgy of healthy development is chosen as parent and carries out seed breeding, parent population and prelarva sample acquire respectively
Fin ray and full fish, are stored in spare in dehydrated alcohol, are operated using Magen animal DNA extracts kit, detect quality and dense
After degree, it is diluted to 100ng/ μ L, -20 DEG C is deposited in and saves backup；
(2) polymorphic micro-satellite markers screening and primer synthesis
8 pairs of micro-satellite primers are chosen, are divided into 3 groups according to clip size, wherein primer AL15, AL20, AL51 are one group,
Primer AL01, AL37, AL18 are one group；FAM, tri- kinds of different fluorophors of HEX, TAMRA are used respectively at the end of forward primer 5 '
It is modified, is analyzed for PCR, primer AL14, AL49 are one group, use FAM, two kinds of differences of HEX respectively at the end of forward primer 5 '
Fluorophor modified, for PCR analyze；
(3) microsatellite locus Genotyping
PCR amplification is carried out to parent and daughter DNA sample using Fluorescence PCR, amplified production is according to institute in step (2)
The method for selecting micro-satellite primers to be grouped is mixed, the sample as upper machine testing；
(4) sample carries out parting on ABI3730XL Genetic Analyser, uses GS-500 as internal reference, uses GeneMapper
3.2 software of V reads the genotype of each sample, is carried out using 3.0 software of CERVUS to parent genotype and progeny genotypes
Analysis, determines the Parent of offspring individual.
The above-mentioned third purpose of the present invention is achieved through the following technical solutions: above-mentioned yellowfin spine porgy microsatellite marker
Application of the specific primer in yellowfin spine porgy paternity test.
Compared with prior art, the present invention has the advantage that
(1) it is flat to establish paternity test using the microsateilite markers of fluorescent marker on yellowfin spine porgy for the first time by the present invention
Platform, identification accuracy rate have reached 99% or more；
(2) present invention can quickly and efficiently identify yellowfin spine porgy difference family and source, be the breeding of yellowfin spine porgy, numerous
It grows combo and enhancement releasing assessment provides foundation；
(3) present invention is combined according to microsatellite clip size and the color difference of fluorescent marker, and 2 or 3 micro- are defended
Star locus products mix sample detection, improve 2-3 times of efficiency than regular-PCR detection method, while having saved cost
And the time；
(4) the microsatellite locus allele number that the present invention selects is more, and polymorphism is high, can be used for yellowfin spine porgy
Population genetic variations, genetic thremmatology assessment and paternity test etc., while can significantly save experimental cost；
(5) method that the present invention uses SSR marker, establishes the paternity test molecular method of yellowfin spine porgy for the first time, and at
Function is applied in the paternity test of yellowfin spine porgy family, is formed and identifies that accurate, cheap SSR paternity test molecule is sentenced
Other group, a kind of convenient, efficient technical method is provided for the fine-variety breeding of yellowfin spine porgy and family management, promotes yellowfin spine
The sound development of porgy industry.
Specific embodiment
Below in conjunction with specific implementation method, the present invention is described in detail.
The screening and synthesis of the specific primer of 1 yellowfin spine porgy microsatellite marker of embodiment
The sequence containing microsatellite marker is selected from yellowfin spine porgy transcript profile library and carrys out design primer, is surveyed by transcript profile
Sequence screens the unigene containing yellowfin spine porgy microsatellite repetitive sequence, then will wherein examine from yellowfin spine porgy transcript profile library
The sequence design microsatellite specific primer containing microsatellite locus is measured, its polymorphism is detected, the final choosing after screening
Select that 8 pairs of bands are clear, primer of the high primer of polymorphism as Parentage determination, primer has in the raw work bioengineering share in Shanghai
The synthesis of limit company uses tri- kinds of different fluorescent bases of FAM, HEX, TAMRA at the end of forward primer 5 ' of each pair of micro-satellite primers respectively
Group is modified and (is shown in Table 1), and primer is synthesized in Shanghai Sheng Gong bioengineering limited liability company.
Table 1 is used for the micro-satellite primers information of yellowfin spine porgy paternity test
2 yellowfin spine porgy microsatellite paternity test method of embodiment
(1) foundation of yellowfin spine porgy parent and descendant relations
112 tail of this experiment yellowfin spine porgy parent population derives from the fish wild seed that Yangjiang city sea is caught, and cultivates in China Water
Hainan Tropical aquatic products research and development centre of South Sea aquatic products research institute of obstetrics research institute Experimental Base.In 393 odd amount in addition to the round numbers generation, derives from
The yellowfin spine porgy mixed family that artificial insemination in December, 2017 obtains.Parent and filial generation sample acquire fin ray and full fish respectively, protect
It is stored in spare in dehydrated alcohol.
(2) extraction of yellowfin spine porgy parent and progeny genome DNA
Using Magen animal tissue DNA extraction kit, 20-50mg tissue sample is processed into fragment small as far as possible, and
It is transferred in 1.5mL centrifuge tube.550 μ L Buffer MTL and 20 μ L Proteinase Ks are added, is vortexed and mixes.55 DEG C of incubated under agitation 3h
Or it is digested overnight sample.5 μ L RNase Solution are added into digestive juice, are mixed by inversion.It is stored at room temperature 30-60min digestion
RNA.13000xg is centrifuged 3min.Supernatant is shifted into new 2.0mL centrifuge tube.500 μ L Buffer DL are added to digestive juice
In, it is vortexed and mixes 20s, 70 DEG C of water-bath 10min.500 μ L dehydrated alcohols are added into digestive juice, is vortexed and mixes 20s.Hipure
GDNA Mini Column is in 2mL collecting pipe.Mixed liquor is shifted into pillar, 10000xg is centrifuged 1min.Pour out outflow
Liquid is reinstalled pillar in collecting pipe, is added on 500 μ L BufferGW1 to pillar, and 10000xg is centrifuged 1min.Pour out stream
Liquid out is reinstalled pillar in collecting pipe, is added on 650 μ L Buffer GW2 to pillar, and 10000xg is centrifuged 1min.It pours out
Efflux is reinstalled pillar in collecting pipe, and 10000xg is centrifuged 2min.By pillar in new 1.5mL centrifuge tube.Add
Enter 30-200 μ L and is preheated to 55 DEG C of Buffer AE to the film center of pillar.2min is placed, 10000xg is centrifuged 1min.Abandon DNA
Each DNA sample is diluted to by column with NanoDrop ND-1000 UV spectrophotometer measuring DNA concentration and quality
100ng/ μ L, DNA save -20 DEG C it is spare.
(3) polymorphic micro-satellite markers screening and reaction system
The sequence containing microsatellite marker is selected from yellowfin spine porgy transcript profile library and carrys out design primer, after screening
Have finally chosen that 8 pairs of bands are clear, primer of the high primer of polymorphism as Parentage determination.
Each pair of micro-satellite primers forward primer 5 ' end respectively with tri- kinds of different fluorophors of FAM, HEX, TAMRA into
Row modification (being shown in Table 1), primer are synthesized in Shanghai Sheng Gong bioengineering limited liability company.
Every group of PCR reaction system total volume is 10 μ L: 1 μ L of distilled water 7.6 μ L, 10 × PCR (TaKaRa) buffer (contains
Mg2+), 0.15 μ L, rTaqDNA polymerase (TaKaRa) of 10mmol/L dNTPs 0.15 μ L, forward and reverse each 0.3 μ L of primer,
0.5 μ L of 100ng/ μ L genomic DNA.
PCR reaction condition are as follows: 95 DEG C of initial denaturation 4min；95 DEG C of denaturation 30s, 60 DEG C of 30s of annealing temperature, 72 DEG C of extensions
40min, 30 circulations；72 DEG C of extension 10min；4 DEG C of preservations.Tm value is shown in Table 1.
(4) microsatellite locus Genotyping and paternity test
Amplified production carries out parting on ABI3730XL Genetic Analyser, uses GS-500LIZ as internal reference, uses
3.2 software of GeneMapper V reads the genotype of individual.Parent and filial generation is calculated using software CERVUS3.0 micro- to defend each
Gene frequency, heterozygosity, expectation heterozygosity, polymorphism information content, average probability of exclusion, Hardy- on constellation position
Weinberg balance and amorph frequency (being shown in Table 2).
The genetic diversity of 28 microsatellite locus of table counts and probability of exclusion
Note: k is allele number, and Ho is observation heterozygosity, HEIt is expected that heterozygosity, PIC are polymorphic content, Excl1
Elimination factor when unknown for parents, elimination factor when Excl2 is known single parent, HW are hardy weinberg equilibrium inspection, and ND expression is not done
It examines, * * * indicates that deviation is extremely significant, and NS indicates that deviation is not significant, and F (Null) indicates amorph frequency.
(5) paternity test result
In the case that parents' gender is unknown, pass through 10000 filial generations of simulation and 112 couples of parents, when confidence level is 95%, 8
The theoretical identification rate of a microsatellite locus is not assigned to Parent successfully up to 99.1%, only 4 offspring individuals.According to yellowfin
The actual gene typing data of spine porgy filial generation has 353 odd amount in addition to the round numbers generation that can find parent when confidence level is 95%.Therefore, of the invention
Practical identification rate be 89.31%.
Therefore, the specific primer of the yellowfin spine porgy microsatellite marker in the present invention can be applied in yellowfin spine porgy parent-offspring
In identification.
Obviously, above content is simply to illustrate that the features of the present invention, and not limitation of the present invention, related technology neck
The those of ordinary skill in domain should belong to protection category of the invention in the variation that corresponding technical field is made according to the present invention.
Sequence table
<110>Nanhai Aquatic Inst., Chinese Aquatic Scientific Research Inst
<120>specific primer of yellowfin spine porgy microsatellite marker and application
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<170> SIPOSequenceListing 1.0
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<213>yellowfin spine porgy (Acanthopagrus latus)
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gctatgctat gacaggcaac c 21
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<213>yellowfin spine porgy (Acanthopagrus latus)
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atttgtggtt ttgatgggga 20
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<213>yellowfin spine porgy (Acanthopagrus latus)
<400> 4
cgtgtgttat tgcctcatgg 20
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<211> 20
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<213>yellowfin spine porgy (Acanthopagrus latus)
<400> 5
caaacgcaga cagcgataag 20
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<213>yellowfin spine porgy (Acanthopagrus latus)
<400> 6
ccacctcaga acccatcagt 20
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<213>yellowfin spine porgy (Acanthopagrus latus)
<400> 7
ttcaacatgt gcggcacg 18
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tattgccctg cacagtgctc cc 22
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cctgggcttt gatatgcct 19
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cagcaacatg ctgccattac 20
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<213>yellowfin spine porgy (Acanthopagrus latus)
<400> 14
gtgtgccctc ataggcagtt 20
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<213>yellowfin spine porgy (Acanthopagrus latus)
<400> 15
cggtagcatt ttcacggtct 20
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<212> DNA
<213>yellowfin spine porgy (Acanthopagrus latus)
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ctgcgagttc acctttcaca 20
Claims (5)
1. a kind of specific primer of yellowfin spine porgy microsatellite marker, it is characterized in that: the specific primer includes 8 pairs of primers,
Respectively AL15, AL20, AL51, AL01, AL37, AL18, AL14 and AL49, in which:
The nucleotide sequence of primer pair AL15 is as shown in SEQ ID NO:1 and SEQ ID NO:2；
The nucleotide sequence of primer pair AL20 is as shown in SEQ ID NO:3 and SEQ ID NO:4；
The nucleotide sequence of primer pair AL51 is as shown in SEQ ID NO:5 and SEQ ID NO:6；
The nucleotide sequence of primer pair AL01 is as shown in SEQ ID NO:7 and SEQ ID NO:8；
The nucleotide sequence of primer pair AL37 is as shown in SEQ ID NO:9 and SEQ ID NO:10；
The nucleotide sequence of primer pair AL18 is as shown in SEQ ID NO:11 and SEQ ID NO:12；
The nucleotide sequence of primer pair AL14 is as shown in SEQ ID NO:13 and SEQ ID NO:14；
The nucleotide sequence of primer pair AL49 is as shown in SEQ ID NO:15 and SEQ ID NO:16.
2. a kind of yellowfin spine porgy microsatellite paternity test method, comprising the following steps:
(1) yellowfin spine porgy DNA is extracted: being acquired yellowfin spine porgy parental animal fin ray and the full fish of filial generation to be identified, is extracted genome
DNA；
(2) polymorphic micro-satellite markers screening and primer synthesis: screening and synthesize 8 pairs of primers in claim 1, and described 8 pairs
Primer is divided into three groups, wherein group one includes AL15, AL20 and AL51, group two includes AL01, AL37 and AL18, and group three includes
AL14 and AL49, group one and group two 5 ' ends of forward primer use respectively the different fluorophor of tri- kinds of FAM, HEX and TAMRA into
Row modification, group three is modified with the two different fluorophors of FAM and HEX respectively at 5 ' ends of forward primer, for PCR points
Analysis；
(3) microsatellite locus Genotyping: three groups of primer pair steps (1) being utilized respectively using Fluorescence PCR in step (2)
In genomic DNA carry out PCR amplification, then amplified production is mixed；
(4) Genotyping is carried out to mixed amplified production, using genotypic results to parent genotype and filial generation gene
Type is analyzed, and determines the Parent of offspring individual.
3. yellowfin spine porgy microsatellite paternity test method according to claim 2, it is characterized in that: fluorescent PCR is anti-in step (3)
At once, every group of reaction system total volume is 10 μ L, comprising: 7.6 μ L, 10 × PCR buffer of distilled water, 1 μ L (contains Mg2+), concentration is
0.15 μ L, rTaqDNA polymerase of dNTPs, the 0.15 μ L of 10mmol/L, forward and reverse each 0.3 μ L of primer, concentration is 100ng/ μ L's
0.5 μ L of genomic DNA.
4. yellowfin spine porgy microsatellite paternity test method according to claim 2, it is characterized in that: fluorescent PCR is anti-in step (3)
At once, PCR reaction condition are as follows: 95 DEG C of initial denaturation 4min；95 DEG C of denaturation 30s, annealing temperature 60 DEG C of 30s, 72 DEG C of extension 40min,
30 circulations；72 DEG C of extension 10min；4 DEG C of preservations.
5. application of the specific primer of yellowfin spine porgy microsatellite marker described in claim 1 in yellowfin spine porgy paternity test.
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| CN111394478A (en) * | 2020-04-27 | 2020-07-10 | 西安理工大学 | PCR (polymerase chain reaction) microsatellite primer and method for paternity test of large yellow croaker by using same |
| CN112899282A (en) * | 2021-03-03 | 2021-06-04 | 中国水产科学研究院南海水产研究所 | SNP (single nucleotide polymorphism) site related to growth traits of yellow fin acanthopagrus latus, and screening method and application thereof |
| CN113667760A (en) * | 2021-07-06 | 2021-11-19 | 中山大学 | SSR marker primer and method for evaluating genetic diversity of sparus latus population |
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| CN111394478B (en) * | 2020-04-27 | 2022-06-17 | 西安理工大学 | PCR (polymerase chain reaction) microsatellite primer and method for paternity test of large yellow croaker by using same |
| CN111394478A (en) * | 2020-04-27 | 2020-07-10 | 西安理工大学 | PCR (polymerase chain reaction) microsatellite primer and method for paternity test of large yellow croaker by using same |
| CN112899282A (en) * | 2021-03-03 | 2021-06-04 | 中国水产科学研究院南海水产研究所 | SNP (single nucleotide polymorphism) site related to growth traits of yellow fin acanthopagrus latus, and screening method and application thereof |
| CN113667760B (en) * | 2021-07-06 | 2023-07-14 | 中山大学 | SSR (simple sequence repeat) marker primer and method for evaluating genetic diversity of sparus praecox population |
| CN113667760A (en) * | 2021-07-06 | 2021-11-19 | 中山大学 | SSR marker primer and method for evaluating genetic diversity of sparus latus population |
| CN114592070A (en) * | 2022-03-07 | 2022-06-07 | 珠海市现代农业发展中心(珠海市金湾区台湾农民创业园管理委员会、珠海市农渔业科研与推广中心) | Method for identifying microsatellite families of sparus latus and application of sparus latus |
| CN114592070B (en) * | 2022-03-07 | 2023-10-17 | 珠海市现代农业发展中心(珠海市金湾区台湾农民创业园管理委员会、珠海市农渔业科研与推广中心) | Microsatellite family identification method and application of sparus flavescens |
| CN115820868B (en) * | 2022-07-27 | 2023-08-29 | 中国水产科学研究院南海水产研究所 | Polymorphic primer for amplifying SNP molecular markers of oplegnathus ruticosus and application thereof |
| CN115820868A (en) * | 2022-07-27 | 2023-03-21 | 中国水产科学研究院南海水产研究所 | Polymorphic primer for amplifying acanthopagrus schlegelii SNP molecular marker and application thereof |
| CN115927662A (en) * | 2022-09-26 | 2023-04-07 | 中国水产科学研究院南海水产研究所 | Amplification primer of bicinchus punctatus SNP locus marker combination and application thereof |
| CN115927662B (en) * | 2022-09-26 | 2024-01-05 | 中国水产科学研究院南海水产研究所 | Amplification primer of SNP locus marker combination of oplegnathus fasciatus and application thereof |
| CN116555433A (en) * | 2022-12-28 | 2023-08-08 | 中国水产科学研究院南海水产研究所 | Primer and kit for identifying Huang Qiji and flat sea bream and application |
| CN116555433B (en) * | 2022-12-28 | 2024-03-19 | 中国水产科学研究院南海水产研究所 | Primer and kit for identifying Huang Qiji and flat sea bream and application |
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