CN108300793B - Microsatellite DNA marker of chinchilla, amplification primer, detection method and application thereof - Google Patents
Microsatellite DNA marker of chinchilla, amplification primer, detection method and application thereof Download PDFInfo
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Abstract
The invention discloses a microsatellite DNA marker of a chinchilla, an amplification primer, a detection method and application thereof. According to the invention, 8 microsatellite markers chrom1-26, chrom2-2, chrom15-6, chrom16-6, chrom16-12, chrom20-9, chrom20-37 and chrom-16 with rich polymorphism are determined by constructing enrichment libraries of mice microsatellite (CT) n, (AG) n, (TG) n and (AC) n, screening and sequencing of positive clones containing microsatellite sequences, and the nucleotide sequences of the clones are respectively shown as SEQ ID No. 1-8. The microsatellite DNA marker of the chinchilla can be applied to the research on the genetic diversity of different groups of the chinchilla and the research on the paternity relationship identification, has good repeatability and is a reliable and effective molecular marker.
Description
The technical field is as follows:
the invention belongs to the technical field of molecular markers, and particularly relates to a microsatellite DNA marker of a chinchilla, an amplification primer, a detection method and application thereof.
Background art:
microsatellite DNA is widely present in prokaryotic and eukaryotic genomes, commonly has two, three and four nucleotide repetitive sequences which account for 50 percent of the eukaryotic genomes, and consists of simple sequence repetitive arrays (such as AC, CCA or GATA and SSR) of 2-6bp in series. Most common are dinucleotide repeats, such as (AC/TG) n and (AG/TC) n. The microsatellite DNA of each specific site consists of two parts: a central core region and peripheral flanking regions. Different individuals in an organism exhibit the same flanking region but different numbers of tandem repeats; it is also possible to have the same number of repeat units but different sizes of flanking regions, or both. DNA RFLP studies of non-microsatellite DNA sites have shown that only 1 to several alleles can be detected by RFLP at each site. Therefore, it is believed that the number of flanking regions of a microsatellite DNA locus is very small, and that the allelic differences at a particular microsatellite locus should be mainly derived from different numbers of tandem repeats.
Microsatellite markers have been widely used in population genetics research, genetic relationship identification, genetic linkage map construction and QTL analysis due to high mutability, co-dominant expression and universality in eukaryotic genomes. Compared with other common nuclear DNA molecular markers (including RFLP, RAPD, AFLP and the like) and allelic enzyme markers, the microsatellite locus serving as a genetic marker has the following advantages: firstly, as a highly polymorphic molecular marker, the microsatellite DNA has the characteristics of high abundance, codominance marker and neutral selection; the microsatellite adopts a single-site DNA fingerprint technology, so that the detection is easy and the repeatability is good; the microsatellite DNA enlarges the sampling range, and reduces the difficulty of sampling work and the influence on the research object; the appearance of the microsatellite provides unprecedented and abundant genetic information data for population biology and promotes the development of corresponding statistical analysis methods.
The yellow chinchilla is a wild species of rat of the rat genus (Rattus) of the family rodentia, is one of the most important pest rats in south China and south-east Asia rice production areas, and is mainly distributed in the south of Yangtze river, the north of lake, Anhui and the like. The chinchilla is in great amount in farmlands of plains, hills and mountainous areas, and is favored by weeds in rice fields, sugarcane fields, vegetable fields, shrubs, pond edges and ditch edges. The yellow chinchilla is a rat with food, is favored by rice, sweet potatoes, wheat and the like, and has serious harm to early-maturing rice. In the absence of vegetarian food, the mice also feed on small animals and insects. The yellow-haired rats are bred at the peak periods of 3 months, 4 months and two periods of 8 months, 9 months and 10 months in one year (Yuanfang and Liu Si Song, 1997). The population quantity change of the chinchillas is influenced by food, climate and environmental factors, and also has the function of adjusting the density in the population and other factors.
With the rapid development of molecular biology technology and molecular genetics, the advantages of molecular markers as individual recognition tools are gradually highlighted, and the molecular markers become one of the analysis means in the aspects of genetic diversity detection, population structure, species interspecies relationship and the like. The yellow chinchilla is one of the main bandicoots in south China, and the inter-population communication and the mating system of the yellow chinchilla have not been studied systematically and deeply. The research on the molecular ecology of the chinchilla is not reported in domestic research at present, the research on the aspect of foreign countries is relatively less, and in order to further understand the information on the aspects of the population structure, the genetic diversity and the like of the chinchilla, a new molecular genetic marker needs to be developed. The research is to research the microsatellite loci of the chinchillas at present, and research the geographic genetic difference of the chinchillas population by using the microsatellite loci, thereby providing a research basis for the future discussion of the relevant fields of the chinchillas and even rodents.
The invention content is as follows:
the invention aims to overcome the defects in the prior art and provides a microsatellite DNA marker of a chinchilla, an amplification primer, a detection method and application thereof. The invention establishes a technology system of the microsatellite DNA of the chinchilla and utilizes the molecular markers to carry out kinship identification and genetic diversity analysis of the chinchilla.
According to the invention, 8 microsatellite markers chrom1-26, chrom2-2, chrom15-6, chrom16-6, chrom16-12, chrom20-9, chrom20-37 and chrom-16 with rich polymorphism are determined by constructing enrichment libraries of luteinis microsatellite (CT) n, (AG) n, (TG) n and (AC) n, screening and sequencing positive clones containing microsatellite sequences.
Therefore, the first object of the invention is to provide a microsatellite DNA marker of a chinchilla, wherein the microsatellite DNA markers are numbered as chrom1-26, chrom2-2, chrom15-6, chrom16-6, chrom16-12, chrom20-9, chrom20-37 and chrom-16;
the nucleotide sequence of chrom1-26 is shown in SEQ ID NO. 1;
the nucleotide sequence of the chrom2-2 is shown as SEQ ID NO. 2;
the nucleotide sequence of chrom15-6 is shown in SEQ ID NO. 3;
the nucleotide sequence of chrom16-6 is shown in SEQ ID NO. 4;
the nucleotide sequence of chrom16-12 is shown in SEQ ID NO. 5;
the nucleotide sequence of chrom20-9 is shown in SEQ ID NO. 6;
the nucleotide sequence of chrom20-37 is shown in SEQ ID NO. 7;
the nucleotide sequence of the chrom-16 is shown as SEQ ID NO. 8.
The second purpose of the invention is to provide an amplification primer marked by the microsatellite DNA of the mice, wherein the amplification primer comprises:
for the chrom1-26 site:
chrom 1-26F: 5'-AGACAATCCAATCTGGGTGC-3' (shown in SEQ ID NO. 9),
chrom 1-26R: 5'-AGGCAATTCTCCACTAGGCA-3' (shown in SEQ ID NO. 10);
for the chrom2-2 site:
chrom 2-2F: 5'-GAAAGCCACTTGGAGGTACG-3' (shown in SEQ ID NO. 11),
chrom 2-2R: 5'-TGCCCAAACAGTGCACTAAG-3' (shown in SEQ ID NO. 12);
for the chrom15-6 site:
chrom 15-6F: 5'-CACTACATCTTCATGCCGGA-3' (shown in SEQ ID NO. 13),
chrom 15-6R: 5'-TCATGGATCATCGTGAAGGA-3' (shown in SEQ ID NO. 14);
for the chrom16-6 site:
chrom 16-6F: 5'-CAGTCTTAGCCCTTGGTGGA-3' (shown in SEQ ID NO. 15),
chrom 16-6R: 5'-CCTCATGTTGTACCGTCCCT-3' (shown in SEQ ID NO. 16);
for the chrom16-12 site:
chrom 16-12F: 5'-TGCTGTTAGCTCCATGTTGG-3' (shown in SEQ ID NO. 17),
chrom 16-12R: 5'-AGGGGTCAGAGTGGTCCTTT-3' (shown in SEQ ID NO. 18);
for the chrom20-9 site:
chrom 20-9F: 5'-GTTTGGCAATTTCCCTCTGA-3' (shown in SEQ ID NO. 19),
chrom 20-9R: 5'-AGACCCCAAACTCTGGCTTT-3' (shown in SEQ ID NO. 20);
for chrom20-37 site:
chrom 20-37F: 5'-AGCTTTCCCTCCCACTCATT-3' (shown in SEQ ID NO. 21),
chrom 20-37R: 5'-TTTTTCACTGCCCAATCACA-3' (shown in SEQ ID NO. 22);
for the chrom-16 site:
chrom-16F: 5'-AAGCGACGCCCATAAACATA-3' (shown in SEQ ID NO. 23),
chrom-16R: 5'-TTCACCCAGGACTTCTTGCT-3' (shown in SEQ ID NO. 24).
The 5' end of the forward primer of the amplification primer is provided with a fluorescent label.
The fluorescent label is preferably FAM, HEX or TAMRA.
The third purpose of the invention is to provide a method for detecting a microsatellite DNA marker of a chinchilla, which comprises the following steps:
(1) extracting the genome DNA of the chinchilla;
(2) taking the genomic DNA extracted in the step (1) as a template, respectively using the primer pair chrom 1-26F and chrom 1-26R aiming at the chrom1-26 site, the primer pair chrom 2-2F and chrom 2-2R aiming at the chrom2-2 site, and the primer pair chrom 15-6F and chrom 15-6R aiming at the chrom15-6 site, carrying out PCR amplification on a primer pair chrom 16-6F and chrom 16-6R aiming at a chrom16-6 site, a primer pair chrom 16-12F and chrom 16-12R aiming at a chrom16-12 site, a primer pair chrom 20-9F and chrom 20-9R aiming at a chrom20-9 site, a primer pair chrom 20-37F and chrom 20-37R aiming at a chrom20-37 site, and a primer pair chrom-16F and chrom-16R aiming at a chrom-16 site;
(3) and typing the PCR amplification product by using a sequencer.
The reaction system of the PCR amplification is preferably 50 mu L, and comprises: 10 XKOD Buffer 5. mu.L, 2mM dNTPs 4. mu.L, 25mM MgSO4mu.L, 100. mu.M forward primer 1.5. mu.L, 100. mu.M reverse primer 1.5. mu. L, DNA template 1. mu. L, KOD-Plus-1. mu.L, water to 50. mu.L.
The reaction procedure of the PCR amplification is preferably as follows: at 95 ℃ for 3min, at 95 ℃ for 30s, at 60 ℃ for 30s, at 68 ℃ for 8min, for 35 cycles; storing at 4 ℃.
The fourth purpose of the invention is to provide the application of the microsatellite DNA marker or the amplification primer in the research of the genetic diversity of the mice population or the paternity test.
The microsatellite DNA marker of the chinchilla can be applied to the research on the genetic diversity of different groups of the chinchilla and the research on the paternity relationship identification, has good repeatability and is a reliable and effective molecular marker.
Description of the drawings:
FIG. 1 is an agarose gel electrophoresis of a genomic DNA of a yellow-haired mouse; wherein, M is DL 2000 Marker, and the strip size from the bottom up is in proper order: 100. 250, 500, 750, 1000 and 2000 bp.
FIG. 2 is an electrophoretogram of a portion of microsatellite locus primers amplified 22 grease mouse samples; wherein 15-6 is an electrophoresis image of 22 chinchilla samples amplified by a chrom15-6 site primer, 16-6 is an electrophoresis image of 22 chinchilla samples amplified by a chrom16-6 site primer, 16-12 is an electrophoresis image of 22 chinchilla samples amplified by a chrom16-12 site primer, and 20-9 is an electrophoresis image of 22 chinchilla samples amplified by a chrom20-9 site primer.
FIG. 3 is SSR typing map of 22 mice genome DNA amplified by chrom1-26 site primer; wherein S1-S22 represent 22 samples of a chinchilla.
FIG. 4 is a SSR typing map of 22 mice genome DNA amplified by chrom2-2 site primer; wherein S1-S22 represent 22 samples of a chinchilla.
FIG. 5 is SSR typing map of 22 mice genome DNA amplified by chrom15-6 site primer; wherein S1-S22 represent 22 samples of a chinchilla.
FIG. 6 is SSR typing map of 22 mice genome DNA amplified by chrom16-6 site primer; wherein S1-S22 represent 22 samples of a chinchilla.
FIG. 7 is a SSR typing map of 22 mice genome DNA amplified by using chrom16-12 site primers; wherein S1-S22 represent 22 samples of a chinchilla.
FIG. 8 is SSR typing map of 22 mice genome DNA amplified by chrom20-9 site primer; wherein S1-S22 represent 22 samples of a chinchilla.
FIG. 9 is a SSR typing map of 22 mice genome DNA amplified by chrom20-37 site primer; wherein S1-S22 represent 22 samples of a chinchilla.
FIG. 10 is a SSR typing map of 22 mice genome DNA amplified by a chrom-16 site primer; wherein S1-S22 represent 22 samples of a chinchilla.
The specific implementation mode is as follows:
the technical solution of the present invention will be further specifically described below by way of specific examples. The practice of the present invention is not limited to the following examples, and any variations and/or modifications of the present invention are intended to fall within the scope of the present invention.
In the invention, the parts and the percentages are weight units, and the adopted equipment and raw materials are all commonly purchased in the field. The methods in the following examples are conventional in the art unless otherwise specified.
Example 1:
1. experimental procedure
1.1, DNA extraction from mice with yellow hair
Approximately 50mg of the muscle of the hamsters is cut and used for extracting DNA, and the extraction method adopts a standard chloroform-isoamylol-alcohol extraction procedure. The purity and concentration of DNA were checked by agarose gel electrophoresis and UV spectrophotometer, and diluted to 30 ng/. mu.L working solution, which was kept at 4 ℃ until use. The DNA concentration was checked by electrophoresis on a 1% agarose gel (see FIG. 1).
1.2 microsatellite locus selection
And (3) referring to the kindred species microsatellite locus of the chinchilla, and searching the SSR locus by utilizing misa software. On both sides of the SSR site, 60 pairs of primers were designed using Primer 3 software.
1.3 primer screening
The initial screening work of 60 pairs of primers is carried out by using a template with better quality, and an amplification system and a reaction program are as follows:
an amplification system:
reaction procedure:
1.4 Synthesis and PCR amplification of fluorescent primers
The forward primer of the primer with better screening effect is subjected to fluorescence labeling, and the synthesized fluorescent primer is used for carrying out PCR amplification on all samples, wherein the amplification system and the amplification conditions are the same as above.
1.5 capillary electrophoresis of fluorescent PCR products
And (3) carrying out gel electrophoresis detection on the PCR product amplified by the fluorescent primer, carrying out capillary electrophoresis according to the detection result, and analyzing the result.
2. Results and analysis of the experiments
2.1 primer prescreening work
According to the designed primers, 60 pairs are selected and synthesized for primer primary screening, 22 chinchilla DNA samples are used for primer primary screening, and 25 pairs of primers are screened.
2.2 fluorescent primer Synthesis amplification and detection
The effectiveness of the 25 pairs of primers screened out initially needs to be further verified, synthesis of fluorescent primers is carried out according to the primers screened out initially, electrophoresis detection is carried out after fluorescent PCR amplification is carried out on all 22 samples, capillary electrophoresis is carried out on the primers screened out again according to the detection result, and partial electrophoresis detection results are shown in figure 2.
2.3 capillary electrophoresis of fluorescent primers
After the amplification product is processed, the amplification product and the internal reference are detected on an ABI 3730 XL machine together to obtain capillary electrophoresis original data, and the identification of bands is carried out by using GeneMaeker software. 8 microsatellite markers with abundant polymorphism are determined (see figures 3-10), and the sites are respectively: chrom1-26 (shown as SEQ ID NO. 1), chrom2-2 (shown as SEQ ID NO. 2), chrom15-6 (shown as SEQ ID NO. 3), chrom16-6 (shown as SEQ ID NO. 4), chrom16-12 (shown as SEQ ID NO. 5), chrom20-9 (shown as SEQ ID NO. 6), chrom20-37 (shown as SEQ ID NO. 7) and chrom-16 (shown as SEQ ID NO. 8).
2.4 Final primer sequences
The sequences of the 8 pairs of primers finally used for genetic clustering analysis are shown in Table 1 below:
TABLE 1 microsatellite markers and amplification primers for mice with yellow hair
2.5 Harder Weinberg equilibrium analysis
The results are given in table 2 below:
TABLE 2 microsatellite marker characteristics table for hamsters
Note: na represents the number of alleles; ne represents the number of effective alleles; ho represents observed heterozygosity values; he represents the desired heterozygosity value; x2Represents a chi-squared check value; the magnitude of the P value indicates significance (P)<0.05 significance)
As can be seen from Table 2, the microsatellite markers chrom1-26, chrom2-2, chrom15-6, chrom16-6, chrom16-12, chrom20-9, chrom20-37 and chrom-16 all have high polymorphism and can be applied to the research of genetic diversity of different populations of the chinchillas and the research of paternity relationship identification.
Sequence listing
<110> institute for biological resource application in Guangdong province
<120> luteinis mouse microsatellite DNA marker and amplification primer, detection method and application thereof
<160> 24
<170> SIPOSequenceListing 1.0
<210> 1
<211> 251
<212> DNA
<213> yellow hair mouse (Rattus losea)
<400> 1
agacaatcca atctgggtgc ctgaacgttt gactcgcaaa ctgcctcctg ctattcgtga 60
agatgagact acgaatacta ctactactac tggaaatgat gatccaggtt gattcactca 120
cgctgtgggc tatcgccaga tcctggccag tacttatgcc agttcatagc aattctactg 180
ttttgccaac ctttttctcc acatcctgtt tgcgtgatgt tccttgtata gtgcctagtg 240
gagaattgcc t 251
<210> 2
<211> 221
<212> DNA
<213> yellow hair mouse (Rattus losea)
<400> 2
gaaagccact tggaggtacg taagtatgag tgctggaaag cgctgctgaa tgtgtgcatg 60
cgtgtgtgtg tgtgtgtgtg tgtgtgtgtg tgtgtgtgtg tgtgtgtgtg aattaagtcc 120
tataatctgt gttttttggt accagtttag tgtcttggtg cactgtctgg gcagaaggat 180
tcctgccctg tgcacgagag tcttagtgca ctgtttgggc a 221
<210> 3
<211> 238
<212> DNA
<213> yellow hair mouse (Rattus losea)
<400> 3
cactacatct tcatgccgga atggtgattc gcgtagatgt tcttgtattg ttgagaattt 60
tgtgtgcctt ttctttcttt ctttctttct ttctttcttt ctttctttct ttcttccttt 120
ctttctttct tttctgtgaa gaattgagtt gcattttgat ggagattttt taatctggag 180
attgctcttg tttagatgac cttttttttt aaccatgctc cttcacgatg atccatga 238
<210> 4
<211> 238
<212> DNA
<213> yellow hair mouse (Rattus losea)
<400> 4
cagtcttagc ccttggtgga aaaccctgga agcacagtgg aaaaacacca ggtgcttaag 60
tttatggtct caggataaaa actcaggact tagcagttgc tacattgtat gctacagcct 120
tgggcaacaa actagcacat cccagggtgt gtgtgtgtgt gtgtgtgtgt gtgtgtgtgt 180
gtgtgtgtgt gtgtggtggg gtggtgagaa cacatgacag ggacggtaca acatgagg 238
<210> 5
<211> 250
<212> DNA
<213> yellow hair mouse (Rattus losea)
<400> 5
tgctgttagc tccatgttgg tttgggctac agtgtgagac tttacctcca ttaaaagctc 60
acctagttag tagttgtgct gttagacaat gaatatattg tctaaaaggc atatataata 120
ataataataa ttatgatagt aacttataaa ccttaaacaa cctttgatgc tcacctgctc 180
cccactcccc agaaaaaaag agaaattgca ctaactgaca aaaccattaa aaaggaccac 240
<210> 6
<211> 196
<212> DNA
<213> yellow hair mouse (Rattus losea)
<400> 6
gtttggcaat ttccctctga attccacatc tgtgctgcag tcagaagatg gctgggaaca 60
catgaaaaaa gacagacaga cagacacgca catacactca cacacacaca tgcacacagg 120
aatgcacgta tgtacacagg cacatgctcg cttgcacagt aatgtaataa aatttaaaag 180
ccagagtttg gggtct 196
<210> 7
<211> 267
<212> DNA
<213> yellow hair mouse (Rattus losea)
<400> 7
agctttccct cccactcatt gccaatcagt gtcctggtgt cccctcaccc tgcctctgtc 60
tctgcaacct gccagcctcc aactgaacag acttccattc ctgtgcaatc taagtcagtc 120
tctccagtct cttcctccct ccctccctcc ctcgctctcc ctctctctta taaaggaaag 180
aaagcactca ctgggtagaa ctgatgtcta tatgcaggtg agggcaggta caagataagg 240
caagacctgt gattgggcag tgaaaaa 267
<210> 8
<211> 230
<212> DNA
<213> yellow hair mouse (Rattus losea)
<400> 8
aagcgacgcc cataaacata taagaagcct acaaaactcc aaatagactg gatcagaaaa 60
gaaaaccctt ctaccacgta ataatcaaaa caccaaatgt acaaacaaca acaacaacat 120
caatattaaa gggactaaga gaaaaaaggt caagcgacat ataaaggcaa acatcagaat 180
aataccaaat gtctcaacag acactatgaa agcaagaagt cctgggtgaa 230
<210> 9
<211> 20
<212> DNA
<213> yellow hair mouse (Rattus losea)
<400> 9
<210> 10
<211> 20
<212> DNA
<213> yellow hair mouse (Rattus losea)
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<210> 11
<211> 20
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<213> yellow hair mouse (Rattus losea)
<400> 11
<210> 12
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<213> yellow hair mouse (Rattus losea)
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<210> 13
<211> 20
<212> DNA
<213> yellow hair mouse (Rattus losea)
<400> 13
<210> 14
<211> 20
<212> DNA
<213> yellow hair mouse (Rattus losea)
<400> 14
<210> 15
<211> 20
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<213> yellow hair mouse (Rattus losea)
<400> 15
<210> 16
<211> 20
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<213> yellow hair mouse (Rattus losea)
<400> 16
<210> 17
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<213> yellow hair mouse (Rattus losea)
<400> 17
<210> 18
<211> 20
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<213> yellow hair mouse (Rattus losea)
<400> 18
<210> 19
<211> 20
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<213> yellow hair mouse (Rattus losea)
<400> 19
<210> 20
<211> 20
<212> DNA
<213> yellow hair mouse (Rattus losea)
<400> 20
<210> 21
<211> 20
<212> DNA
<213> yellow hair mouse (Rattus losea)
<400> 21
<210> 22
<211> 20
<212> DNA
<213> yellow hair mouse (Rattus losea)
<400> 22
<210> 23
<211> 20
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<213> yellow hair mouse (Rattus losea)
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Claims (7)
1. A little satellite DNA marker combination of mice of yellow hair used for the study of the genetic diversity of mice colony or paternity test research, characterized by, the said little satellite DNA marker is numbered as chrom1-26, chrom2-2, chrom15-6, chrom16-6, chrom16-12, chrom20-9, chrom20-37 and chrom-16;
the nucleotide sequence of chrom1-26 is shown in SEQ ID NO. 1;
the nucleotide sequence of the chrom2-2 is shown as SEQ ID NO. 2;
the nucleotide sequence of chrom15-6 is shown in SEQ ID NO. 3;
the nucleotide sequence of chrom16-6 is shown in SEQ ID NO. 4;
the nucleotide sequence of chrom16-12 is shown in SEQ ID NO. 5;
the nucleotide sequence of chrom20-9 is shown in SEQ ID NO. 6;
the nucleotide sequence of chrom20-37 is shown in SEQ ID NO. 7;
the nucleotide sequence of the chrom-16 is shown as SEQ ID NO. 8.
2. An amplification primer of the luteus microsatellite DNA marker combination of claim 1, wherein said amplification primer is as follows:
for the chrom1-26 site:
chrom1-26 F：5’-AGACAATCCAATCTGGGTGC-3’，
chrom1-26 R：5’-AGGCAATTCTCCACTAGGCA-3’；
for the chrom2-2 site:
chrom2-2 F：5’-GAAAGCCACTTGGAGGTACG-3’，
chrom2-2 R：5’-TGCCCAAACAGTGCACTAAG-3’；
for the chrom15-6 site:
chrom15-6 F：5’-CACTACATCTTCATGCCGGA-3’，
chrom15-6 R：5’-TCATGGATCATCGTGAAGGA-3’；
for the chrom16-6 site:
chrom16-6 F：5’-CAGTCTTAGCCCTTGGTGGA-3’，
chrom16-6 R：5’-CCTCATGTTGTACCGTCCCT-3’；
for the chrom16-12 site:
chrom16-12 F：5’-TGCTGTTAGCTCCATGTTGG-3’，
chrom16-12 R：5’-AGGGGTCAGAGTGGTCCTTT-3’；
for the chrom20-9 site:
chrom20-9 F：5’-GTTTGGCAATTTCCCTCTGA-3’，
chrom20-9 R：5’-AGACCCCAAACTCTGGCTTT-3’；
for chrom20-37 site:
chrom20-37 F：5’-AGCTTTCCCTCCCACTCATT-3’，
chrom20-37 R：5’-TTTTTCACTGCCCAATCACA-3’；
for the chrom-16 site:
chrom-16 F：5’-AAGCGACGCCCATAAACATA-3’，
chrom-16 R：5’-TTCACCCAGGACTTCTTGCT-3’；
the 5' end of the forward primer of the amplification primer is provided with a fluorescent label.
3. The amplification primer of claim 2, wherein the fluorescent label is FAM, HEX or TAMRA.
4. A method for detecting a microsatellite DNA marker of a chinchilla is used for the research of the genetic diversity of the chinchilla population or the identification research of the paternity relationship, and comprises the following steps:
(1) extracting the genome DNA of the chinchilla;
(2) using the genomic DNA extracted in step (1) as a template, using the primer pair chrom 1-26F and chrom 1-26R for chrom1-26 site, the primer pair chrom 2-2F and chrom 2-2R for chrom2-2 site, and the primer pair chrom 15-6F and chrom 15-6R for chrom15-6 site, respectively, as described in claim 3, carrying out PCR amplification on a primer pair chrom 16-6F and chrom 16-6R aiming at a chrom16-6 site, a primer pair chrom 16-12F and chrom 16-12R aiming at a chrom16-12 site, a primer pair chrom 20-9F and chrom 20-9R aiming at a chrom20-9 site, a primer pair chrom 20-37F and chrom 20-37R aiming at a chrom20-37 site, and a primer pair chrom-16F and chrom-16R aiming at a chrom-16 site;
(3) and typing the PCR amplification product by using a sequencer.
5. The detection method according to claim 4, wherein the PCR amplification reaction system is 50 μ L, and comprises: 10 XKOD Buffer 5. mu.L, 2mM dNTPs 4. mu.L, 25mM MgSO4mu.L, 100. mu.M forward primer 1.5. mu.L, 100. mu.M reverse primer 1.5. mu. L, DNA template 1. mu. L, KOD-Plus-1. mu.L, water to 50. mu.L.
6. The detection method according to claim 4 or 5, wherein the PCR amplification is performed by the following reaction procedures: at 95 ℃ for 3min, at 95 ℃ for 30s, at 60 ℃ for 30s, at 68 ℃ for 8min, for 35 cycles; storing at 4 ℃.
7. The use of the amplification primer of claim 2 in a study of the genetic diversity of a cohort of mice of the species hamsters, or in an paternity test.
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