CN113005202A - SNP marker related to salt tolerance of tilapia and application thereof - Google Patents
SNP marker related to salt tolerance of tilapia and application thereof Download PDFInfo
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- CN113005202A CN113005202A CN202110326008.7A CN202110326008A CN113005202A CN 113005202 A CN113005202 A CN 113005202A CN 202110326008 A CN202110326008 A CN 202110326008A CN 113005202 A CN113005202 A CN 113005202A
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- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/124—Animal traits, i.e. production traits, including athletic performance or the like
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/156—Polymorphic or mutational markers
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- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
- Y02A40/80—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in fisheries management
- Y02A40/81—Aquaculture, e.g. of fish
Abstract
The invention belongs to the technical field of aquatic organisms, and discloses an SNP marker related to salt tolerance of tilapia and application thereof. The sequence of the SNP marker is shown as SEQ ID NO: 1 or SEQ ID NO: 4, the sequence shown in SEQ ID NO: 2942bp of 1 sequence or SEQ ID NO: the SNP site base at 99bp of the 4 sequence is G or T. The invention creatively provides an SNP marker related to the salt tolerance of tilapia, on the basis, the SNP marker can be used for screening or detecting salt tolerance individuals of tilapia, is not limited by the growth stage, can greatly improve the breeding process of tilapia and quickly select tilapia germplasm with excellent salt tolerance, develops a primer pair and a kit for corresponding detection on the basis, can quickly judge the genotype of tilapia to be detected, and has great scientific value and commercial value.
Description
Technical Field
The invention belongs to the technical field of aquatic organisms, and particularly relates to an SNP marker related to salt tolerance of tilapia and application thereof.
Background
Tilapia (Tilapia), a widely salty tropical fish of the family of Paciformes (Perciformes) Paciformes (Cichlida), originally produced in Africa, commonly known as African crucian carp. Tilapia mossambica as an important economic fish has the characteristics of fresh and tender meat, high nutritional value, quick growth and the like, is deeply favored by consumers and farmers, is listed as one of six main foods by the Food and Agriculture Organization (FAO) of the United nations, and is cultured in more than 100 countries and regions. China is the largest tilapia producing and consuming country in the world, the yield in 2019 reaches 164.17 ten thousand tons, which accounts for about 60% of the total world tilapia yield, and the tilapia mossambica, nile tilapia, oreochromis aureus, hybrid species and other breeding varieties are mainly used, and the salt tolerance of different breeding varieties is greatly different.
The shortage of global water resources and the warming of climate make available fresh water resources sharply reduced, and the fresh water aquaculture industry faces serious water resource shortage. With the enhancement of the protection of the fresh water environment in China, the area of available water areas for fresh water culture is in short supply, and the fresh water culture industry of tilapia is greatly restricted. The saline-alkali water resource in China is quite abundant and occupies about 55 percent of the total area of lakes in China, but the saline-alkali water has high carbonate salinity (HCO)3 -And CO3 2-) The high pH value and the complex ion composition are not beneficial to the growth of conventional fish species, the development and the utilization of saline-alkali water areas have great limitations, the utilization rate of the saline-alkali water resources in most areas is still very low, and the development and the utilization of the saline-alkali water resources are very urgent and necessary. At present, the utilization of salt-tolerant germplasm resources is an important means for solving the problem. Tilapia mossambica is euryhaline fish, and can grow and reproduce in saline water or seawater through domestication and breeding. Therefore, developing research on the development of salt-tolerant molecular markers of tilapia and cultivating a salt-tolerant tilapia variety suitable for being cultured in saline water or seawater have important significance for the development and utilization of the water areas and the sustainable development of fishery.
The breeding of the salt-tolerant tilapia by using the traditional breeding means has the problems of low selection rate, long period and the like. Molecular marker assisted breeding is an effective means for solving the problem. Single Nucleotide Polymorphisms (SNP) are one of the most commonly used molecular marking methods, refer to the change of DNA sequences caused by the change of a Single base, and have the characteristics of large quantity, wide distribution, high genetic stability, high positioning precision, simple typing operation, simple detection method, low requirements on large-scale experiments and detection automation and the like. Due to the advantages of high whole genome coverage rate, high genotyping efficiency and the like, the method is widely applied to correlation analysis and molecular assisted breeding. The molecular marker assisted breeding technology is a modern breeding technology for indirectly selecting target traits through molecular markers closely linked with the target traits, has the characteristics of high selection strength, no environmental influence, reliable result and the like, can realize early seed selection and improve the accuracy of breeding so as to obtain greater genetic progress and faster breeding process.
Disclosure of Invention
Aiming at the defects and shortcomings of the prior art, the invention aims to provide an SNP marker related to the salt tolerance of tilapia.
The invention also aims to provide a primer pair or a kit for detecting the SNP marker related to the salt tolerance of tilapia.
The invention further aims to provide application of the kit in screening or detecting salt-tolerant tilapia parents.
The purpose of the invention is realized by the following technical scheme:
an SNP marker related to salt tolerance of tilapia, wherein the sequence of the SNP marker is shown as SEQ ID NO: 1 or SEQ ID NO: 4, and the sequence shown in SEQ ID NO: 1 sequence 2942bp or SEQ ID NO: the SNP site base at 99bp of the 4 sequence is G (guanine) or T (thymine).
The SNP marker is SEQ ID NO: 1 or the nucleotide sequence 2942 th base shown as SEQ ID NO: 4 is represented by Y, and Y represents G or T.
Furthermore, the SNP marker is a dominant allele with G base at 2942bp of the SEQ ID NO.1 sequence or 99bp of the SEQ ID NO.4 sequence, and when tilapia individuals only contain a homozygous genotype with G as the SNP site (namely GG genotype), the individuals are selected for salt-tolerant targets.
Further, the SNP marker may include the above gene sequence, for example, the sequence of SEQ ID NO.1 including G at 2942bp as described above; or, for example, also the sequence amplified by the sequence SEQ ID NO.2 and the sequence SEQ ID NO.3 (sequence SEQ ID NO. 4), such a sequence being the sequence SEQ ID NO.4 itself or the sequence SEQ ID NO.1 including the sequence SEQ ID NO. 4.
Still further, the SNP marker may also include the sequence of SEQ ID NO.1 itself which is T at 2942 bp.
The applicant researches and discovers that a G (guanine) or T (thymine) type SNP exists at 2942bp of a tilapia TRPV4 gene sequence SEQ ID NO.1, is located at the 3' end of a non-coding region of a gene, and is named as a TRPV4-G2942T SNP site.
The applicant further researches and discovers that experiments on salt stress of tilapia show that the SNP site at 2942bp in the TRPV4 gene has obvious correlation with salt tolerance of tilapia, and the individual salt tolerance of the site corresponding to GG genotype is obviously stronger than that of GT genotype or TT genotype.
A primer pair for detecting the SNP marker related to the salt tolerance of tilapia is disclosed, and the nucleotide sequence of the primer pair is as follows:
primer 1: 5'-AGAAAGCCATCAGCATCAGT-3' (SEQ ID NO: 2);
primer 2: 5'-AGTGTCTCTGCATGTCGTCT-3' (SEQ ID NO: 3).
The primer pair can be used for amplifying the SNP marker or a sequence containing the marker, or detecting whether the tilapia with the SNP marker exists or not, so as to further screen and/or detect the salt-tolerant tilapia, or further identify and/or screen the salt-tolerant tilapia parent.
A kit for detecting the SNP marker related to the salt tolerance of tilapia comprises the primer pair.
The kit comprises a reagent for detecting the SNP marker, such a reagent can comprise, for example, a PCR amplification reagent including a primer pair, a sample DNA extraction reagent and the like, and such a reagent can also be considered as any reagent and combination thereof for realizing the method for screening and/or detecting the salt-tolerant tilapia or parents.
The kit is applied to screening or detecting salt-tolerant tilapia parents.
Through detecting the genotype corresponding to the SNP marker of the tilapia, salt-tolerant tilapia can be effectively screened and/or detected, and the tilapia which is detected to obtain the polymorphic site of the base at 2942bp of the sequence of the SEQ ID NO.1 and has the GG genotype is salt-tolerant tilapia or a tilapia parent.
In addition, SNP is a molecular marker that can be inherited from parents to progeny, and parents containing SNP markers related to a desired trait can be selected and propagated to obtain progeny containing the trait, thereby accelerating the breeding process.
Preferred GG genotypes in the present invention are homozygotes that do not segregate in progeny inheritance. Therefore, the SNP marker can be used for breeding new salt-tolerant tilapia species by screening/detecting, tilapia parents can be selected according to the genotype of candidate parents on the SNP marker site, and the tilapia parents containing GG genotypes are selected for pairing propagation, so that offspring fries with strong salt tolerance can be obtained, the new tilapia species suitable for saline water or seawater culture can be quickened, the culture cost can be reduced, and the breeding time can be saved.
Further, the application method comprises the following steps:
extracting tilapia DNA by adopting SEQ ID NO: 2 sequence and SEQ ID NO: 3, and selecting a primer pair with the sequence shown in SEQ ID NO: tilapia mossambica of which the genotype corresponding to the polymorphic site of the base at 2942bp of the sequence 1 is GG genotype or the genotype corresponding to the polymorphic site of the base at 99bp of the sequence SEQ ID NO.4 is GG genotype is taken as salt-tolerant tilapia mossambica or parent.
Specifically, for example, when the SNP site base polymorphic site of a tilapia individual is detected to be only G, namely the genotype corresponding to the marker site is GG genotype, the detection sample is judged to be salt-tolerant tilapia and can be selected as a breeding parent of the salt-tolerant tilapia.
Further, the PCR amplification reaction system is a 25 μ L reaction system, and is specifically as follows:
further, the PCR amplification reaction conditions are as follows: pre-denaturation at 94 ℃ for 2 min; denaturation at 94 ℃ for 30 seconds, annealing at 54 ℃ for 20 seconds, extension at 72 ℃ for 30 seconds, 32 cycles; extension at 72 ℃ for 7 min.
Compared with the prior art, the invention has the beneficial effects that:
(1) the invention creatively provides an SNP marker related to tolerance of tilapia, and confirms the correlation between different genotypes and salt tolerance of tilapia, on the basis, the SNP marker can be used for screening or detecting tilapia, or screening or detecting salt tolerance tilapia or parents thereof, has positive effects in the field of breeding or breeding of tilapia, provides a basis for auxiliary selection breeding of salt tolerance molecular markers of tilapia, provides reliable genetic data for screening and establishing new salt tolerance tilapia strains, and has great scientific value and commercial value.
(2) The primer pair provided by the invention has good specificity and high amplification efficiency.
Detailed Description
The present invention will be described in further detail with reference to examples, but the embodiments of the present invention are not limited thereto.
Example 1
The function research of the SNP marker of the invention comprises the following steps:
the Holana dragon tilapia (Oreochromys hornorum) (500 tails) used by the invention is from the high-grade aquatic germplasm base of the Zhujiang aquatic research institute of Chinese aquatic science. Salinity stress experiments were performed in fish ponds at 320cm x 200cm x 150 cm. The culture is carried out for 7 days before the experiment, during which the dissolved oxygen is 8.7 +/-0.6 mg/L, the water temperature is 28.5 +/-1 ℃, and the pH value is 8.4 +/-0.5. Selecting healthy and active tilapia mossambica to perform an experiment, adding pre-prepared saline water about 113 per mill to increase the salinity of pool water every 30 minutes after the experiment begins, increasing the salinity by 5 per mill each time, and detecting the actual salinity of the pool water by using a handheld salinity meter. During the period, the air is continuously inflated, and no feed is fed. And (3) beginning to die tilapia after 2 hours, recording indexes of death time, water salinity, body quality and the like of each tilapia, and storing the fins in absolute ethyl alcohol. Under the condition of salt stress, 100 individuals died first are considered as a salt-sensitive group, the salt tolerance is low, and 100 individuals died last are considered as a salt-tolerant group, so that the salt tolerance is high. Genomic DNA was extracted and the allele and genotype frequencies of the SNP site at 2942bp of SEQ ID NO.1 were examined are shown in Table 1.
TABLE 1 Oreochromis mossambicus TRPV4-G2942T SNP locus allele and genotype frequency
From the above results, it can be seen that the genotype frequency of the TRPV4-G2942T SNP site GG in the salt tolerance group is 92%, while the genotype of TT in the early death salt sensitive group is 77%, so that the SNP site is considered to be closely related to the salt tolerance of tilapia mossambica, and the genotype corresponding to the site GG is the salt tolerance genotype.
Therefore, the TRPV4-G2942T SNP (2942 bp of SEQ ID NO.1) molecular marker obtained by the invention can be used for assisting in breeding tilapia salt-tolerant varieties, when only G is detected on the site of an individual to be tested by tilapia, namely the corresponding genotype of the individual is GG genotype, the individual is tested to be a salt-tolerant individual and can be selected as a parent of the tilapia salt-tolerant varieties, and the TRPV4-G2942T SNP (2942 bp of SEQ ID NO.1) site is not only G but also T, or only T, namely the corresponding genotype of the individual is GT or TT genotype, the individual is a salt-sensitive tilapia individual.
Example 2
This example screens/detects SNP markers of tilapia:
(first) extraction of sample DNA
(1) Taking 3mg of fin ray tissue of tilapia sample to be detected, shearing, adding 0.5mL of lysis solution (10mmol/L Tris-HCl; 0.1mol/L EDTA; 0.5% SDS; 30mg/L RNase; 100mg/L proteinase K, pH8.0), digesting for 1 hour at 55 ℃;
(2) adding a phenol/chloroform/isoamyl alcohol (volume ratio is 25: 24: 1) mixture with the same volume, uniformly mixing, standing at room temperature for 5 minutes, centrifuging at 12000 r/min for 10 minutes, taking a supernatant, extracting with chloroform once, standing at room temperature for 5 minutes, centrifuging at 12000 r/min for 10 minutes, and taking the supernatant;
(3) adding 2 times volume of absolute ethyl alcohol, standing at room temperature for 10 minutes to precipitate DNA, and centrifuging at 12000 r/min for 10 minutes to obtain precipitate;
(4) the precipitate was washed with 70% ethanol 1 time, centrifuged at 12000 rpm for 2 minutes, the supernatant was aspirated, allowed to stand at room temperature for 10 minutes, and dissolved in 50. mu.l of TE (10mmol/L Tris-HCl; 1mmol/L EDTA, pH8.0) to obtain a DNA solution which was stored at 4 ℃ for further use.
(II) design and Synthesis of primers
Designing and synthesizing a pair of primers according to a tilapia TRPV4 gene sequence (SEQ ID NO.1) for amplifying SNP loci at 2942bp of SEQ ID NO.1 or 99bp of SEQ ID NO.4, wherein the primer sequences are as follows:
primer 1: 5'-AGAAAGCCATCAGCATCAGT-3' (SEQ ID NO. 2);
primer 2: 5'-AGTGTCTCTGCATGTCGTCT-3' (SEQ ID NO. 3).
(III) PCR reaction:
reaction System (25. mu.L System)
The PCR reaction conditions are as follows: pre-denaturation at 94 ℃ for 2 min; denaturation at 94 ℃ for 30 seconds, annealing at 54 ℃ for 20 seconds, extension at 72 ℃ for 30 seconds, 32 cycles; extension at 72 ℃ for 7 min.
(IV) genotype analysis (screening/detection) of Tilapia mossambica
And (2) sequencing the PCR amplification product (namely the sequence of SEQ ID NO. 4) obtained in the step (three), checking a sequencing peak image, analyzing the genotype of each tilapia, and according to the comparison result of the sequence of SEQ ID NO.1 or the sequence of SEQ ID NO.4 with the measured sequence, determining that individuals with G single peak at 2942pb of the sequence of SEQ ID NO.1 are GG genotype individuals, individuals with GT double peaks are GT genotype individuals, and individuals with T single peak are TT genotype individuals. Wherein GG genotype individuals are screened/detected as salt-tolerant tilapia mossambica or tilapia mossambica parents, and GT or TT genotype individuals are screened/detected as salt-sensitive tilapia mossambica individuals.
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.
Sequence listing
<110> Zhujiang aquatic research institute of Chinese aquatic science research institute
<120> SNP marker related to salt tolerance of tilapia and application thereof
<130> 2021-3-17
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aggagcgcac agtagatcag agaaaacatg tccaaacttt ctatgggata aagttacgga 60
gcaggctgta aacatttcta agaaaatgaa aactggattt actggttaga ggaagcgccc 120
acatgcttca gcatcatttc cagtttcccc ccgaagagga atttggcaca tcattgtgat 180
aagagggatg aatgaggggc gatctgctat cttcaaaagg cgccacctcg ccttgcccaa 240
gggcaacgcc atcagctctg aaccaagcat ctcggtggat ttgggagaca gtgaggctgc 300
tcagcctgag ggtgatggag ccttcccttt atctgagttc tcacacctgt tcgagagcca 360
ggatggatcc ccagcaactc aggactcgag tcaggagtca atcctggagc cggcccagcc 420
gggccacccc gcagacagca gacagtacct acggatgaag ttccacggtg ctttcaagaa 480
gggcatatcc aacccaatgg acctcctgga atccaccata tatgagtcaa atgtggttcc 540
tgctcccaag aaagcaccca tggactcact ctttgactat ggcacctacg gaaactcaag 600
caaccagaag aaacgcagaa agaagctccc aaaaggcaaa acagaggcat cctgtgacga 660
gagccagagc tctgatcctc caaaagtcgt caaagtattc aaccgctcgc ttctcttcga 720
ctgtgtgtca cgtggagacc ccggggaact cgagggcctg ttggagtacc tgcaaagtaa 780
caacaagagg ttaaccgatg aggaatttag agagccatac acaggtaaga catgtctgcc 840
aaaagctttg atgaaccttt atggccgtca gaacaacacc atcccagtgc tggtcgacat 900
agcagagaag aatgggagtc tcagagagtt cattaatacg cccttcaggg acgtctacta 960
cagaggccag acagcgctcc acattgccat cgaacgacgc tgtaagcagt atgtgaagct 1020
gctggtggaa aaaggagctg acgttcacgc ccaggcgagg ggacgcttct tccagcccaa 1080
agacgagggt ggttacttct actttggtga gctgcctctc tctctggctg cctgcacaaa 1140
ccagcccgac atagtgcact acctgacaga gaatccgcac aagaaagctg acgtgcgacg 1200
ccaggattca cgtggaaaca cggtgctgca cgcgctggtg cacattgcag acaacaccaa 1260
ggacaacacg cgtttcctca caaagatgta cgacctgctg ctaatcaaga ctgccaagct 1320
ttacccagac tgcaacctgg agacagtgcc caacaatgat ggcatgtcgc ctctcatgat 1380
ggctgccaga ttgggcaaga tagggatttt tcaacacatt atccgacgtg agatcaaaga 1440
cgaggaagtt cgtcatctgt cccgtaagtt taaggactgg gcctatggtc cggtgtactc 1500
ctccctctat gatctgtctt cactggatac atgtggcaag gagtcatctg tgctggaaat 1560
ccttgtctac accagtcaca atgagaaccg ccatgagatg ctggcagtgg agcccatcaa 1620
tgagctgttg agggccaaat ggaacaggtt tgctgctgtc actttttaca tcagtgtgtt 1680
ctcctacctc atcaccatga tcatattcac cctggtggct tattaccaac caacagatgg 1740
aaagcctcca tacccacata ccacatcgtc tgactactgg cggatggccg gggagattgt 1800
cacactggca tcaggaatct tcttcttcct cacaaatatt aaggacctct tcctgaaaaa 1860
gtgccctggg gtgaagtctt tatttattga tggatccttt caactgctgt acttcatcta 1920
ctctgtactg attatagtca cagctgctct ctacctgtct ggcattgagg cctacgtctc 1980
tgtcatggtg tttgcactcg ccctgggctg gatgaacacg ctttacttca ccaggggcct 2040
gaagctcacg ggcacctaca gcatcatgat acaaaagatt cttttcaagg accttttcag 2100
gtttctgctg gtgtatgtgc tcttcatgat cggttttgca tcagccttgg tatccctgct 2160
gactgtgtgc cctccgccag gcacagtgtg taacgggagt tgcccaacct accctgcatg 2220
cagggacaat aacactttca gcgctttcct tctggacctc ttcaagctca ccattgggat 2280
gggagacctg gacatgatct acagtgcaca gaatccagtc gtctttctca tcctgttagt 2340
tacttacatc attctcacct tcgtgctgct actcaacatg ttgatcgctt tgatggggga 2400
gacggtgggt caggtctcca aggagagcaa aaaaatctgg aagcttcagt gggcaacgac 2460
tatcttggac atcgagcgct cgttcccggt ctgtcttcgc aagtcttttc gagttggaga 2520
gatggtgacg gtggggaaga actatgatgg cacacctgat cggcgctggt gtttcagggt 2580
ggacgaggtt aactggtgtc actggaatca gaacctggcg ataattaacg aggatccggg 2640
caagagtgag accatccaag ccaacgggct gcagcagggt gttagagctt tgaggagaga 2700
tcgctggtcc acagtggtcc cgcgggcggt ggagttgagc aaaggttctc agtctcatga 2760
tctcgcggta gagatggagc cgctgtcacc caggcactga ttcccgcaga tgatgagtgc 2820
tgtccttgtt atgaggagac agcagaaagc catcagcatc agtgaaatca ctctgcaggg 2880
aacgatttgg ccaataagtg tactaatttg cacgcgtctc ctgtgtcttt tgttcagaaa 2940
ayctgtcacg aatataaaag cagcacactc tgtgtaacag gacagacgac atgcagagac 3000
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caaaaaagga taacaaaaaa tgtgttttgt aagcagtgag ctgtaatttg ttttgtactt 3120
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agaaagccat cagcatcagt gaaatcactc tgcagggaac gatttggcca ataagtgtac 60
taatttgcac gcgtctcctg tgtcttttgt tcagaaaayc tgtcacgaat ataaaagcag 120
cacactctgt gtaacaggac agacgacatg cagagacact 160
Claims (8)
1. An SNP marker related to salt tolerance of tilapia, which is characterized in that: the sequence of the SNP marker is shown as SEQ ID NO: 1 or SEQ ID NO: 4, and the sequence shown in SEQ ID NO: 1 sequence 2942bp or SEQ ID NO: the SNP site base at 99bp of the 4 sequence is G or T.
2. A primer pair for detecting SNP markers related to salt tolerance of tilapia according to claim 1, wherein the nucleotide sequences of the primer pair are as follows:
primer 1: 5'-AGAAAGCCATCAGCATCAGT-3' (SEQ ID NO: 2);
primer 2: 5'-AGTGTCTCTGCATGTCGTCT-3' (SEQ ID NO: 3).
3. A kit for detecting the SNP marker related to the salt tolerance of tilapia according to claim 1, which is characterized in that: comprising the primer pair of claim 2.
4. The use of the kit of claim 3 in screening or detecting salt-tolerant tilapia parents.
5. The application of the kit according to claim 4 in screening or detecting salt-tolerant tilapia parents, which is characterized in that: detecting the genotype corresponding to the SNP marker of the tilapia mossambica to obtain the position 2942bp of the sequence of SEQ ID NO.1 or the sequence of SEQ ID NO: the tilapia with the genotype of GG corresponding to the polymorphism site of the base at 99bp of the 4 sequence is salt-tolerant tilapia or a tilapia parent.
6. The application of the kit according to claim 4 or 5 in screening or detecting salt-tolerant tilapia parents is characterized in that the application method comprises the following steps:
extracting tilapia DNA by adopting SEQ ID NO: 2 sequence and SEQ ID NO: 3, and selecting a primer pair with the sequence shown in SEQ ID NO: the tilapia with the genotype of GG corresponding to the polymorphic site of the base at 2942bp of the sequence 1 or the polymorphic site of the base at 99bp of the sequence SEQ ID NO.4 is used as the salt-tolerant tilapia or parent.
7. The application of the kit according to claim 6 in screening or detecting salt-tolerant tilapia parents, which is characterized in that: the PCR amplification reaction system is a 25 mu L reaction system, and comprises the following specific steps:
8. the application of the kit according to claim 6 in screening or detecting salt-tolerant tilapia parents, which is characterized in that: the PCR amplification reaction conditions are as follows: pre-denaturation at 94 ℃ for 2 min; denaturation at 94 ℃ for 30 seconds, annealing at 54 ℃ for 20 seconds, extension at 72 ℃ for 30 seconds, 32 cycles; extension at 72 ℃ for 7 min.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110326008.7A CN113005202B (en) | 2021-03-26 | 2021-03-26 | SNP marker related to salt tolerance of tilapia and application thereof |
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Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20030047143A1 (en) * | 2001-08-27 | 2003-03-13 | Streelman Jeffrey Todd | Method for Identifying Fast- Growing Fish |
| US20090165156A1 (en) * | 2006-06-20 | 2009-06-25 | Norwegian School Of Veterinary School | Method for increasing the saltwater tolerance of a fish |
| CN106916884A (en) * | 2016-12-26 | 2017-07-04 | 中山大学 | A kind of bolti salt tolerant related molecular marker SSR450 and its application |
| CN107217094A (en) * | 2017-06-14 | 2017-09-29 | 海南华大海洋科技有限公司 | One SNP marker related to the gift tilapia speed of growth and its application |
| WO2018214187A1 (en) * | 2017-05-23 | 2018-11-29 | 中国科学院南海海洋研究所 | Snp marker related to low-salt tolerance characteristic of litopenaeus vannamei, amplification primer therefor, and application thereof |
| CN110241230A (en) * | 2019-06-26 | 2019-09-17 | 中国水产科学研究院淡水渔业研究中心 | Identify method, kit and the application of Oreochromis aureus, bolti and its hybrid generation |
| CN111690755A (en) * | 2020-06-24 | 2020-09-22 | 中国水产科学研究院淡水渔业研究中心 | Marker, method, kit and application for improving nile tilapia breeding efficiency |
-
2021
- 2021-03-26 CN CN202110326008.7A patent/CN113005202B/en active Active
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20030047143A1 (en) * | 2001-08-27 | 2003-03-13 | Streelman Jeffrey Todd | Method for Identifying Fast- Growing Fish |
| US20090165156A1 (en) * | 2006-06-20 | 2009-06-25 | Norwegian School Of Veterinary School | Method for increasing the saltwater tolerance of a fish |
| CN106916884A (en) * | 2016-12-26 | 2017-07-04 | 中山大学 | A kind of bolti salt tolerant related molecular marker SSR450 and its application |
| WO2018214187A1 (en) * | 2017-05-23 | 2018-11-29 | 中国科学院南海海洋研究所 | Snp marker related to low-salt tolerance characteristic of litopenaeus vannamei, amplification primer therefor, and application thereof |
| CN107217094A (en) * | 2017-06-14 | 2017-09-29 | 海南华大海洋科技有限公司 | One SNP marker related to the gift tilapia speed of growth and its application |
| CN110241230A (en) * | 2019-06-26 | 2019-09-17 | 中国水产科学研究院淡水渔业研究中心 | Identify method, kit and the application of Oreochromis aureus, bolti and its hybrid generation |
| CN111690755A (en) * | 2020-06-24 | 2020-09-22 | 中国水产科学研究院淡水渔业研究中心 | Marker, method, kit and application for improving nile tilapia breeding efficiency |
Non-Patent Citations (6)
| Title |
|---|
| DAN LI JIANG 等: "Identifying a Long QTL Cluster Across chrLG18 Associated with Salt Tolerance in Tilapia Using GWAS and QTL-seq", 《MARINE BIOTECHNOLOGY》, vol. 21, no. 2, 30 April 2019 (2019-04-30), pages 250, XP036743146, DOI: 10.1007/s10126-019-09877-y * |
| FENG LIU 等: "A microsatellite-based linkage map of salt tolerant tilapia (Oreochromis mossambicus x Oreochromis spp.) and mapping of sex-determining loci", 《BMC GENOMICS》, vol. 14, no. 58, 28 January 2013 (2013-01-28), pages 1 - 14 * |
| GRACE LIN 等: "Mapping QTL for Sex and Growth Traits in Salt-Tolerant Tilapia (Oreochromis spp. X O. mossambicus)", 《PLOS ONE》, vol. 11, no. 1, 11 November 2016 (2016-11-11), pages 1 - 17 * |
| 刘玉姣 等: "罗非鱼催乳素Ⅰ基因的组织分布及盐胁迫对其表达的影响", 《南方水产科学》, vol. 10, no. 6, 5 December 2014 (2014-12-05), pages 51 - 57 * |
| 杨玲 等: "5个罗非鱼品种(系)线粒体DNA 16S rRNA和Cyt b基因片段序列分析", 《长江大学学报(自然科学版)农学卷》, vol. 5, no. 3, 15 September 2008 (2008-09-15), pages 35 - 41 * |
| 王楠 等: "瓦氏雅罗鱼耐碱性状相关分子标记的筛选", 《中国水产科学》, vol. 11, no. 6, 15 November 2015 (2015-11-15), pages 1105 - 1114 * |
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