A SSR marker of Sargassum semifolium and its amplification primers, detection method and application

Abstract

The present invention discloses a SSR marker of half-leaf Sargassum and its amplification primer, detection method and application. The present invention uses the half-leaf Sargassum sequence obtained by genome survey sequencing to obtain 14 highly polymorphic SSR markers, which are numbered S12, S16, S24, S31, S33, S39, S41, S53, S58, S72, S74, S85, S108 and S143. The SSR marker of the present invention has the characteristics of strong polymorphism and high heterozygosity, and has a strong advantage in the diversity assessment of half-leaf Sargassum population with low genetic diversity. The SSR marker amplification primer and detection method of the present invention can be used for the genetic diversity analysis of half-leaf Sargassum population, and have application prospects in the identification of half-leaf Sargassum population.

Description

Technical Field

[0001] The invention belongs to the technical field of molecular markers, and in particular relates to a Sargassum semifolium SSR marker and an amplification primer, a detection method and application thereof. Background Art

[0002] The ocean is the "blue granary" for humans to obtain high-quality protein. my country's subtropical seas are vast and the habitats are complex and diverse. The coral reef ecosystem is an important ecosystem in the ocean and an important source of seafood to meet people's lives. Sargassum is one of the important members of the coral reef ecosystem. In recent years, with the rapid development of my country's aquaculture industry, the domestic market demand for Sargassum has increased. Therefore, in order to restore the balance of the coral reef ecosystem, it is urgent to carry out artificial transplantation and ecological aquaculture of Sargassum.

[0003] Sargassum hemiphyllum belongs to the genus Sargassum, the order Fucaleso, the family Sargassaceae, the phylum Phaeophyta. It mainly grows near the low tide line and on rocks about one meter deep. It also grows in large rock marshes in the low tide zone. It is a common and dominant species along the coasts of the East China Sea and the South China Sea. The algae is large and grows fast. It is an excellent species for the reconstruction of artificial seaweed fields and a high-quality raw material for the extraction of feed, algin and beverages.

[0004] Germplasm resources are the material basis for the development of aquaculture. The data of germplasm resource evaluation are of great significance for maintaining the genetic diversity of species during artificial transplantation and ecological aquaculture, and provide a theoretical basis for resource conservation and aquaculture effect evaluation. Microsatellites (also known as simple sequence repeats, SSRs) have been widely used in the evaluation of population diversity due to their advantages such as high heterozygosity, rich polymorphism, good repeatability, and co-dominant inheritance. So far, the development of SSR markers for Sargassum has not been carried out. This study hopes to lay a foundation for the study of genetic diversity, population identification, resource conservation, and aquaculture effect evaluation of large algae on islands and reefs in the South China Sea through the development of highly polymorphic SSR markers. Summary of the invention

[0005] The purpose of the present invention is to provide a SSR marker of half-leaf Sargassum and its amplification primers, detection method and application, so as to provide a highly polymorphic molecular marker for genetic diversity analysis and population identification of half-leaf Sargassum.

[0006] To achieve the above-mentioned purpose of the invention, the present invention performs genome survey sequencing on three samples of half-leaf Sargassum collected from the sea area of ​​Xuwen Jiaowei in Zhanjiang and the sea area near Yangmeikeng in Shenzhen, and the obtained sequence is used to search for SSR sites through MISA (MIcroSAtellite identification tool) software, and the SSR sites with geographical differences are screened out using a self-compiled program, and a total of 513 SSR sites are obtained. Primer Premier 5 software is used to randomly design primers for 260 sites among the 513 SSR sites screened out. The requirements for primer design are: primer length 18-22bp, GC content 50-60%, Tm value 58-65℃, the difference in Tm value of upstream and downstream primers is not greater than 5, and primer dimers, hairpin structures and mismatches are avoided as much as possible; the length of the amplified product is 100-550bp. A total of 260 pairs of SSR primers were designed and synthesized in the experiment, and PCR amplification was performed on the genome DNA of half-leaf Sargassum, of which 42 pairs of primers can stably amplify the target band. The 42 screened primer pairs were used to amplify four DNA samples of half-leaf Sargassum from four areas, namely, the Jiaowei waters of Xuwen, Zhanjiang, the waters of Naozhou Island, Zhanjiang, the waters near Yangmeikeng, Shenzhen, and the waters of Wanshan Islands. The PCR amplification products were typed using a 3730XL sequencer, and the length of the allele fragments was read using GeneMapper3.2 software. The polymorphism information was analyzed using Cervus 3.0 software, and finally 14 highly polymorphic SSR markers of half-leaf Sargassum were identified.

[0007] The first object of the present invention is to provide an SSR marker of Sargassum hemiphyllum, wherein the SSR markers are numbered S12, S16, S24, S31, S33, S39, S41, S53, S58, S72, S74, S85, S108 and S143;

[0008] The nucleotide sequence of S12 is shown in SEQ ID NO.1;

[0009] The nucleotide sequence of S16 is shown in SEQ ID NO.2;

[0010] The nucleotide sequence of S24 is shown in SEQ ID NO.3;

[0011] The nucleotide sequence of S31 is shown in SEQ ID NO.4;

[0012] The nucleotide sequence of S33 is shown in SEQ ID NO.5;

[0013] The nucleotide sequence of S39 is shown in SEQ ID NO.6;

[0014] The nucleotide sequence of S41 is shown in SEQ ID NO.7;

[0015] The nucleotide sequence of S53 is shown in SEQ ID NO.8;

[0016] The nucleotide sequence of S58 is shown in SEQ ID NO.9;

[0017] The nucleotide sequence of S72 is shown in SEQ ID NO.10;

[0018] The nucleotide sequence of S74 is shown in SEQ ID NO.11;

[0019] The nucleotide sequence of S85 is shown in SEQ ID NO.12;

[0020] The nucleotide sequence of S108 is shown in SEQ ID NO.13;

[0021] The nucleotide sequence of S143 is shown in SEQ ID NO.14.

[0022] The second object of the present invention is to provide amplification primers for the above-mentioned Sargassum hemiphyllum SSR marker, wherein the amplification primers include:

[0023] Targeting the S12 locus:

[0024] F12: 5'-TTCATTTCAAGCATGTCCCA-3';

[0025] R12: 5'-GTGGATTCCGAATGACGACT-3';

[0026] Targeting the S16 site:

[0027] F16: 5'-GAATAATGGCATCGTGGTCC-3';

[0028] R16: 5'-GACGCGCTCGAATATTTGTT-3';

[0029] Targeting the S24 site:

[0030] F24: 5'-CGTTCCTTGTCGGAACATTT-3';

[0031] R24: 5'-TGCTCTGTGCACCCACTTAC-3';

[0032] Targeting the S31 site:

[0033] F31: 5'-AGTACGCAAACGCTCCCTTA-3';

[0034] R31: 5'-GCTTCGGATGAAAGATCAGC-3';

[0035] Targeting the S33 site:

[0036] F33: 5'-TACAGTTGCGGATAGTGGCA-3';

[0037] R33: 5'-CCCTTCGTCGCGTCTATTA-3';

[0038] Targeting the S39 site:

[0039] F39: 5'-GAAGGACGTGTACCTGGCAT-3';

[0040] R39: 5'-CTCGCACTACAAACAGCAGC-3';

[0041] Targeting the S41 site:

[0042] F41: 5'-GTGACCACGGACTGGATCTT-3';

[0043] R41: 5'-CGAAGCTGCACACAAAGTA-3';

[0044] Targeting the S53 locus:

[0045] F53: 5'-TTCTGATCTTGTGACCACGG-3';

[0046] R53: 5'-CCCGGATAGATGAGGTTTGA-3';

[0047] Targeting the S58 site:

[0048] F58: 5'-TGTGACCACAAGTAGCCTGC-3';

[0049] R58: 5'-CCATGATACCTTTGTCCGCT-3';

[0050] Targeting the S72 site:

[0051] F72: 5'-TGGCGTTACAAAGTGTGAGG-3';

[0052] R72: 5'-CCGGTACCATTTCATTTGCT-3';

[0053] Targeting the S74 site:

[0054] F74: 5'-GGAGAACAGTCGGACACGAT-3';

[0055] R74: 5'-GATCTCCACGGTAGAACCGA-3';

[0056] Targeting the S85 site:

[0057] F85: 5'-CGTCCCTTTGGAACACAGAT-3';

[0058] R85: 5'-TAGCGATCGACACGTCAAAG-3';

[0059] Targeting the S108 site:

[0060] F108: 5'-ACGCTTGCGAAAGGACTATG-3';

[0061] R108: 5'-TAGTTGGGTTGCCGTAGGTC-3';

[0062] Targeting the S143 site:

[0063] F143: 5'-AGTTTGTAATCGTGGGCTGG-3';

[0064] R143: 5'-TGTGACCACAAGTAGCCTGC-3'.

[0065] Preferably, the 5' end of the forward primer of the amplification primer is labeled with a fluorescent group.

[0066] Preferably, the fluorescent group is FAM, HEX or TAMRA.

[0067] The third object of the present invention is to provide a reagent for detecting the above-mentioned SSR markers for use in preparing products for genetic diversity analysis and population identification of Sargassum semi-leaf.

[0068] The fourth object of the present invention is to provide the use of the above-mentioned SSR markers or the above-mentioned amplification primers in the preparation of a kit for genetic diversity analysis and population identification of Sargassum semi-leaf.

[0069] The fifth object of the present invention is to provide a kit for genetic diversity analysis and population identification of Sargassum semifolium, which comprises the above-mentioned amplification primers.

[0070] The sixth object of the present invention is to provide a method for detecting SSR markers of Sargassum hemiphyllum, comprising the following steps:

[0071] (1) Extracting genomic DNA of Sargassum hemiphyllum;

[0072] (2) Using the genomic DNA extracted in step (1) as a template, primers F12 / R12 for S12 site, primers F16 / R16 for S16 site, primers F24 / R24 for S24 site, primers F31 / R31 for S31 site, primers F33 / R33 for S33 site, primers F39 / R39 for S39 site, primers F39 / R39 for S41 site were respectively used. PCR amplification was performed using primers F41 / R41, primers F53 / R53 for S53 site, primers F58 / R58 for S58 site, primers F72 / R72 for S72 site, primers F74 / R74 for S74 site, primers F85 / R85 for S85 site, primers F108 / R108 for S108 site, and primers F143 / R143 for S143 site;

[0073] (3) typing the PCR product amplified in step (2) using a sequencer;

[0074] (4) Perform genetic diversity analysis on the typing results obtained in step (3).

[0075] Preferably, the PCR amplification reaction system is 25 μL and includes: 2+ 10×PCR buffer 2.5μL, 25mM MgCl2 2.0μL, 10mM dNTP 0.5μL, high-fidelity PCR enzyme 1U, 20μM forward primer 1μL, 20μM reverse primer 1μL, DNA template 20ng, and the rest is made up to 25μL with sterile double distilled water.

[0076] Preferably, the PCR amplification has the following reaction procedure: pre-denaturation at 95°C for 5 minutes; denaturation at 95°C for 30 seconds, annealing at 56°C for 30 seconds, extension at 72°C for 30 seconds, for a total of 35 cycles; and extension at 72°C for another 6 minutes.

[0077] The seventh objective of the present invention is to provide the application of the SSR marker, the amplification primer, the kit or the detection method in the genetic diversity analysis and population identification of Sargassum semi-leaf.

[0078] The present invention uses the SSR sequence of half-leaf Sargassum obtained from the genome survey sequencing database to develop 14 highly polymorphic half-leaf Sargassum SSR markers, which are numbered S12, S16, S24, S31, S33, S39, S41, S53, S58, S72, S74, S85, S108 and S143. The half-leaf Sargassum SSR markers of the present invention can be used for genetic diversity analysis and population tracing of half-leaf Sargassum, providing a theoretical basis for genetic diversity research, resource conservation and aquaculture effect evaluation of large algae on South China Sea islands and reefs. BRIEF DESCRIPTION OF THE DRAWINGS

[0079] Figure 1 This is a cluster diagram of Sargassum semi-leaf originating from 4 different regions, among which Xw: Xuwen Jiaowei waters of Zhanjiang, Nz: Naozhou Island waters of Zhanjiang, Sz: waters near Yangmeikeng of Shenzhen, and Ws: Wanshan Islands waters. DETAILED DESCRIPTION

[0080] The present invention is further described below with reference to the embodiments, but is not limited thereto.

[0081] The experimental methods in the following examples, unless otherwise specified, are all conventional methods or carried out according to the instructions of the kit. The materials, reagents, etc. used in the following examples, unless otherwise specified, can be obtained from commercial channels. Primer synthesis and sequencing were completed by Sangon Biotech (Shanghai) Co., Ltd.

[0082] Example 1

[0083] Four samples of Sargassum semi-leaf were collected from the sea area near Yangmeikeng, Shenzhen, the sea area of ​​Wanshan Islands, the sea area of ​​Xuwen Jiaowei, Zhanjiang, and the sea area of ​​Naozhou Island, Zhanjiang, for a total of 16 strains; a commercially available DNA extraction kit was used to extract the genomic DNA of the 16 Sargassum semi-leaf. The extracted DNA was used as a template and 14 pairs of primers (Table 1) were used to perform PCR amplification analysis and capillary electrophoresis typing on the 16 samples. The PCR amplification reaction system included: Mg-free 2+ 10×PCR buffer 2.5μL, 25mMMgCl2 2.0μL, 10mM dNTP 0.5μL, high-fidelity PCR enzyme 1U, 20μM forward primer 1μL, 20μM reverse primer 1μL, DNA template 20ng, the rest is supplemented to 25μL with sterile double distilled water; the reaction program is: 95℃ pre-denaturation for 5 minutes; 95℃ denaturation for 30 seconds, 56℃ annealing for 30 seconds, 72℃ extension for 30 seconds, a total of 35 cycles; 72℃ extension for 6 minutes. The PCR amplification products were typed using a 3730XL sequencer, and the length of the allele fragments was read using GeneMapper 3.2 software. The polymorphism information was analyzed using Cervus 3.0 software. The results are shown in Table 2.

[0084] The number of alleles (Na) of each SSR marker was between 11 and 20, with an average of 14.36 alleles; the number of effective alleles (Ne) was between 7.822 and 13.8, with an average of 10.383 effective alleles; the Shannon diversity index (I) was between 2.459 and 2.768, with an average of 2.62; the observed heterozygosity (Ho) was between 0.507 and 0.981, with an average of 0.731; the expected heterozygosity was between 0.851 and 0.989, with an average of 0.931; the polymorphic information content (PIC) was between 0.811 and 0.937, with an average of 0.893. UPGMA cluster analysis was performed by MEGA to construct a cluster tree such as Figure 1 The results showed that the genetic diversity analysis of 16 strains of Sargassum hemiphyllum from different sources using 14 SSR markers was consistent with the geographical distribution.

[0085] Table 1 14 pairs of primers used in PCR

[0086]

[0087]

[0088] Table 2 Genetic diversity parameters of 14 SSRs in 16 Sargassum hemiphyllum strains from different sources

[0089]

[0090]

[0091] N: number of samples, Na: number of alleles, Ne: effective number of alleles, I: Shannon diversity index, Ho: observed heterozygosity, He: expected heterozygosity, PIC: polymorphic information content, Mean: mean value.

[0092] Example 2

[0093] A total of 95 samples of Sargassum semi-leaf were collected from the sea area of ​​Jiaowei, Xuwen, Zhanjiang, the sea area of ​​Naozhou Island, Zhanjiang, the sea area of ​​Shenzhen and the sea area of ​​Wanshan Islands, including 16 strains (SZ-1) from the sea area near Dameisha, Shenzhen, 16 strains (SZ-2) from the sea area near Yangmeikeng, Shenzhen, 15 strains (SZ-3) from Dapeng Bay, Shenzhen, 16 strains (WS) from the sea area of ​​Wanshan Islands, 16 strains (XW) from the sea area of ​​Jiaowei, Xuwen, Zhanjiang, and 16 strains (NZ) from the sea area of ​​Naozhou Island, Zhanjiang. The DNA of the genome of 95 Sargassum semi-leaf was extracted using a commercially available DNA extraction kit, and the extracted DNA was used as a template, and 14 pairs of primers (Table 1) were used to perform PCR amplification analysis and capillary electrophoresis typing on the above 95 individuals, respectively, and the PCR amplification conditions were the same as those in Example 1. The PCR amplification products were typed using a 3730XL sequencer, the length of the allele fragments was read using GeneMapper 3.2 software, and the polymorphism information was analyzed using Cervus 3.0 software. The results are shown in Table 3.

[0094] In this example, the number of samples of the six natural populations of half-leaf Sargassum is 15 in the SZ-3 population, and the other five populations are all 16 samples; the average number of alleles (Na) is between 12.5 and 13.5, the average number of effective alleles (Ne) is between 7.489 and 8.482, the average Shannon diversity index (I) is between 2.226 and 2.239, the average observed heterozygosity (Ho) is between 0.047 and 0.663, the average expected heterozygosity is between 0.784 and 0.912, and the average polymorphic information content (PIC) is between 0.836 and 0.912.

[0095] Table 3 Genetic diversity parameters in six natural populations of Sargassum hemiphyllum from different sources

[0096]

[0097] N: number of samples, Na: number of alleles, Ne: effective number of alleles, I: Shannon diversity index, Ho: observed heterozygosity, He: expected heterozygosity, Mean: mean, SE: standard error, PIC: polymorphic information content.

[0098] The embodiments described above are only descriptions of the preferred implementation modes of the present invention, and are not intended to limit the scope of the present invention. Without departing from the design spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by ordinary engineering and technical personnel in the field should fall within the protection scope determined by the claims of the present invention. Sequence Listing <110> Institute of South China Sea Oceanology, Chinese Academy of Sciences <120> A SSR marker of Sargassum semifolium and its amplification primers, detection method and application <160> 14 <170> SIPOSequenceList 1.0 <210> 1 <211> 212 <212> DNA <213> Vegetables (Sargassum hemiphyllum) <400> 1 ttcatttcaa gcatgtccca tcatacagta tacgcgatac ttaaaaggag ggcgtatgac gctgccgcct accatacgta tgccctagcc ccatagggtc accatacgta tagtagc attctagtag tagtagtagt tagtagtagtagtagtcaccg gaggagcatt ctatcttcta gtagtcgtca ttcggaatcc ac 212 <210> 2 <211> 216 <212> DNA <213> Vegetables (Sargassum hemiphyllum) <400> 2 gaataatggc atcgtggtcc acggtaccgt ccgcaagctt cttgggcttg atcacctgtg 120. aagcaaagca aagccataaa atactacttt tagcccctct tatcgagagt acatgtgtgt attaggggaa gggagtacgc aaaagaaatt ctgataatag aataataataataataata ctacctccac ttgtacaaca aatattcgag cgcgtc <210> 3 <211> 165 <212> DNA <213> Sargassum hemiphyllum <400> 3 cgttccttgt cggaacattt atctgccgag acaaaaaaaa gcgccagacc agacgaagca 60 gggatgcaga aaaattaaca tggaatggaa tggaatggaa tggaatggaa tggaatggaa 120 cgggataaga taggatagag gcaacgtaag tgggtgcaca gagca 165 <210> 4 <211> 253 <212> DNA <213> Sargassum hemiphyllum <400> 4 agtacgcaaa cgctccctta tattattaat atagtatagc tccgcgtctt cactcgttga 60 aattaaatta aattaaatta aattaaatga aaccatctta aatcttttgg ttaccccgtt 120 cgttcgtcca cgcataacag gctgcagaag ctaaggccat catctcatgg cttatcacag 180 aaggtggtgc agacgtaaat gcggtcgacc tcgccggacg actgcctggt gaggctgatc 240 tttcatccga agc 253 <210> 5 <211> 246 <212> DNA <213> Sargassum hemiphyllum <400> 5 tacagttgcg gatagtggca gtaataatag cagcagcagt agtaatagca gcagcagtag 60 tggcaatagt atagcagca gcagtagtgg cagtaataat ggtagcagca gttgtagtag 120 tagtagtatt tattaagttg cacataaccg cgcaacccgg cccttccaat gctacaggta 180 cggatccacc ctatggaga tcagtagttg aactgtgac aactcctaat aagacgcgac 240 Gagggg 246 <210> 6 <211> 160 <212> DNA <213> Sargassum hemiphyllum <400> 6 gaaggacgtg tacctggcat ggggtatcgg ttctggcgt gcggtttatt gtgcaacttc 60 cccaatgacg ttgtgaccac aagtactttg cggttagtct acgtaagtga gataataat 120 aatataata aaaggact gctgctgttt gtagtgcgag 160 <210> 7 <211> 116 <212> DNA <213> Sargassum hemiphyllum <400> 7 gtgaccacgg actggatctt tgggatcagc taataatgta gcaatcaat caatcaatca 60 atcaatcaat gttgcctgc cttcagcagc caagcgtact ttggtgtgca gcttcg 116 <210> 8 <211> 256 <212> DNA <213> Sargassum hemiphyllum <400> 8 ttctgatctt gtgaccacgg gctggatctt tgagattagc ttatgtgaga attcaatcaa 60 tcaatcaatc aatcaaatca cctattttga tctcaaccga cacgattcta aaacccatga 120 aatacggtct ctgttctttc caagaaacct cgaatatttc gtcagctttg tagtggtgat 180 aaggattgtc tgaaagtcaa ccaatactta aacgtggcaa caaaaaatgt ttgccctcaa 240 acctcatcta tccggg 256 <210> 9 <211> 163 <212> DNA <213> Sargassum hemiphyllum <400> 9 tgtgaccaca agtagcctgc ctttgcggtt agtctacgta agtgagaata ataataataa 60 taataataat aataataata ataacaataa taataatgaa tcttcttcgc gacggtgaca 120 cacgtggcct acacgcgttt cggagcggac aaaggtatca tgg 163 <210> 10 <211> 231 <212> DNA <213> Sargassum hemiphyllum <400> 10 tggcgttaca aagtgtgagg atttacgtat tttatatggc atagaaagaa aatagtgtac 60 gtcgaactgg gaaaaatac caatatccg acatttacct tcagaaaagg gaaagatcga caagaggggcg agaataaaaa agacgccaaa caagtacta ctactactac father atgtgacaga tacgaaggag aggaataat aagcaaatga aatggtaccg g <210> 11 <211> 276 <212> DNA <213> Vegetables (Sargassum hemiphyllum) <400> 11 60. ggagaacagt cggacacgat gagtgcctgc tgtagcggtg ttggtcttac tttgtgtgtc tatgtatgtg cctcgtaatg atttgatttg atccccttgt tcagctgacc acaagcagga 120 ttggcggctg tagtagtgc gcataatgtg ctaaatgtga gtaaacaaca acaacaacaa ccactaccac agatctcgcc acagacgtac agagtgaaaa gggcatatac tacacgtgta cctggcgcac cggacgtcgg ttctaccgtg gagatc 276 <210> 12 <211> 189 <212> DNA <213> Vegetables (Sargassum hemiphyllum) <400> 12 cgtccctttg cgtcccttc cgttgtgttc tattcgag aacgcgtcaa cctatctgca gcagtaagag cagcagcagc agtaagagca gcagaagcag cagcagcagc agcagtagta 120 gtttcctgct tatcactcga taagtatcga cgatcggcgg gaactattac tttgacgtgt 180 cgatcgcta 189 <210> 13 <211> 119 <212> DNA <213> Sargassum hemiphyllum <400> 13 acgcttgcga aaggactatg gccgaaagaa gtattgtatt gtattgtatt gtattgtatt 60 gttcacacgt tcagcgtacg atgtccacga aactgaagag acctacggca acccaacta 119 <210> 14 <211> 264 <212> DNA <213> Sargassum hemiphyllum <400> 14 agtttgtaat cgtgggctgg ttttccactg cctgcttgaa atttgcggag caacatcagt 60 gccagtcgcc tgcgtggacg gtatcgagac atttctgcaa cctaatagta gcaatcggta 120 ccacagctag atatagcagt gggatctcgc gtttgcctct tgggccaacc acacgtctct 180 gttgtgaagc ccttattatt attattatta ttattattct cacttacgta gactaaccgc 240 aaatgcaggc tacttgtggt caca 264

Claims

1. A SSR marker for Sargassum hemiphyllum, characterized in that: The SSR markers are numbered S12, S16, S24, S31, S33, S39, S41, S53, S58, S72, S74, S85, S108 and S143; The primers for amplifying the S12 marker are: F12: 5'-TTCATTTCAAGCATGTCCCA-3'; R12: 5'-GTGGATTCCGAATGACGACT-3'; The primers for amplifying the S16 marker are: F16: 5'-GAATAATGGCATCGTGGTCC-3'; R16: 5'-GACGCGCTCGAATATTTGTT-3'; The primers for amplifying the S24 marker are: F24: 5'-CGTTCCTTGTCGGAACATTT-3'; R24: 5'-TGCTCTGTGCACCCACTTAC-3'; The primers for amplifying the S31 marker are: F31: 5'-AGTACGCAAACGCTCCCTTA-3'; R31: 5'-GCTTCGGATGAAAGATCAGC-3'; The primers for amplifying the S33 marker are: F33: 5'-TACAGTTGCGGATAGTGGCA-3'; R33: 5'-CCCTTCGTCGCGTCTATTA-3'; The primers for amplifying the S39 marker are: F39: 5'-GAAGGACGTGTACCTGGCAT-3'; R39: 5'-CTCGCACTACAAACAGCAGC-3'; The primers for amplifying the S41 marker are: F41: 5'-GTGACCACGGACTGGATCTT-3'; R41: 5'-CGAAGCTGCACACCAAAGTA-3'; Primers for amplifying the S53 marker are: F53: 5'-TTCTGATCTTGTGACCACGG-3'; R53: 5'-CCCGGATAGATGAGGTTTGA-3'; Primers for amplifying the S58 marker are: F58: 5'-TGTGACCACAAGTAGCCTGC-3'; R58: 5'-CCATGATACCTTTGTCCGCT-3'; Primers for amplifying the S72 marker are: F72: 5'-TGGCGTTACAAAGTGTGAGG-3'; R72: 5'-CCGGTACCATTTCATTTGCT-3'; Primers for amplifying the S74 marker are: F74: 5'-GGAGAACAGTCGGACACGAT-3'; R74: 5'-GATCTCCACGGTAGAACCGA-3'; Primers for amplifying the S85 marker are: F85: 5'-CGTCCCTTTGGAACACAGAT-3'; R85: 5'-TAGCGATCGACACGTCAAAG-3'; Primers for amplifying the S108 marker are: F108: 5'-ACGCTTGCGAAAGGACTATG-3'; R108: 5'-TAGTTGGGTTGCCGTAGGTC-3'; The primers for amplifying the S143 marker are: F143: 5'-AGTTTGTAATCGTGGGCTGG-3'; R143: 5'-TGTGACCACAAGTAGCCTGC-3'.

2. An amplification primer for the SSR marker of Sargassum hemiphyllum according to claim 1, characterized in that: The amplification primers include: Targeting the S12 locus: F12: 5'-TTCATTTCAAGCATGTCCCA-3'; R12: 5'-GTGGATTCCGAATGACGACT-3'; Targeting the S16 site: F16: 5'-GAATAATGGCATCGTGGTCC-3'; R16: 5'-GACGCGCTCGAATATTTGTT-3'; Targeting the S24 site: F24: 5'-CGTTCCTTGTCGGAACATTT-3'; R24: 5'-TGCTCTGTGCACCCACTTAC-3'; Targeting the S31 site: F31: 5'-AGTACGCAAACGCTCCCTTA-3'; R31: 5'-GCTTCGGATGAAAGATCAGC-3'; Targeting the S33 site: F33: 5'-TACAGTTGCGGATAGTGGCA-3'; R33: 5'-CCCTTCGTCGCGTCTTATTA-3'; for S39 site: F39: 5'-GAAGGACGTGTACCTGGCAT-3'; R39: 5'-CTCGCACTACAAACAGCAGC-3'; for S41 site: F41: 5'-GTGACCACGGACTGGATCTT-3'; R41: 5'-CGAAGCTGCACACCAAAGTA-3'; for S53 site: F53: 5'-TTCTGATCTTGTGACCACGG-3'; R53: 5'-CCCGGATAGATGAGGTTTGA-3'; for S58 site: F58: 5'-TGTGACCACAAGTAGCCTGC-3'; R58: 5'-CCATGATACCTTTGTCCGCT-3'; Targeting the S72 site: F72: 5'-TGGCGTTACAAAGTGTGAGG-3'; R72: 5'-CCGGTACCATTTCATTTGCT-3'; Targeting the S74 site: F74: 5'-GGAGAACAGTCGGACACGAT-3'; R74: 5'-GATCTCCACGGTAGAACCGA-3'; for S85 site: F85: 5'-CGTCCCTTTGGAACACAGAT-3'; R85: 5'-TAGCGATCGACACGTCAAAG-3'; Targeting the S108 site: F108: 5'-ACGCTTGCGAAAGGACTATG-3'; R108: 5'-TAGTTGGGTTGCCGTAGGTC-3'; Targeting the S143 site: F143: 5'-AGTTTGTAATCGTGGGCTGG-3'; R143: 5'-TGTGACCACAAGTAGCCTGC-3'.

3. The amplification primer according to claim 2, characterized in that The 5' end of the forward primer of the amplification primer is labeled with a fluorescent group.

4. The amplification primer according to claim 3, characterized in that The fluorescent group is FAM, HEX or TAMRA.

5. Use of a reagent for detecting the SSR markers described in claim 1 in preparing a product for analyzing the genetic diversity of a Sargassum semifolium population.

6. Use of the amplification primers according to claim 2 in preparing a kit for analyzing the genetic diversity of Sargassum hemiphyllum population.

7. A kit for analyzing genetic diversity of Sargassum semifolium population, characterized in that: Contains the amplification primers described in claim 2, 3 or 4.

8. A method for detecting SSR markers of Sargassum hemiphyllum, characterized in that: The following steps are involved: (1) Extracting genomic DNA of Sargassum hemiphyllum; (2) using the genomic DNA extracted in step (1) as a template, and performing PCR amplification using the amplification primers described in claim 2, 3 or 4; (3) typing the PCR product amplified in step (2) using a sequencer; (4) Perform genetic diversity analysis on the typing results obtained in step (3).

9. The detection method according to claim 8, characterized in that: The PCR amplification reaction system is 25 μL, including: Mg-free 2+ 10×PCR buffer 2.5μL, 25mM MgCl2 2.0μL, 10mM dNTP 0.5μL, high-fidelity PCR enzyme 1U, 20μM forward primer 1μL, 20μM reverse primer 1μL, DNA template 20ng, and the rest is made up to 25μL with sterile double-distilled water; The reaction procedure was as follows: pre-denaturation at 95°C for 5 minutes; denaturation at 95°C for 30 seconds, annealing at 56°C for 30 seconds, and extension at 72°C for 30 seconds, for a total of 35 cycles; and extension at 72°C for another 6 minutes.

10. Use of the SSR marker described in claim 1, the amplification primer described in claim 2, the kit described in claim 7 or the detection method described in claim 8 in the genetic diversity analysis of Sargassum semifolium population.

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