A specific SNP locus combination for identifying Lingao pig breeds and its application

Abstract

The present invention belongs to the field of molecular biological identification, and specifically relates to a specific SNP site combination and application for identifying the Lingao pig variety. The specific SNP site combination is screened based on a whole genome association analysis combined with a selection signal analysis method, comprising 61 SNP sites, and the physical location is determined by sequence comparison based on the reference genome Sscrofa11.1 of the pig. The specific SNP site combination can accurately identify the unique genetic characteristics of Lingao pigs at the gene level, and can be used for the precise protection of the genetic resources of Hainan Lingao pigs.

Description

Technical Field

[0001] The invention belongs to the field of molecular biological identification, and specifically relates to a specific SNP site combination and application for identifying a Lingao pig breed. Background Art

[0002] Lingao pig is a Hainan specialty pig mainly distributed in Lingao County, Hainan Province. It is a pig breed that is used for both meat and fat, and is one of the important groups of Hainan pigs. Roasted suckling pig made from Lingao suckling pig is a specialty food in Hainan Province, with the characteristics of thin and crispy skin, tender meat, fragrant taste, and crispy bones.

[0003] Although Lingao pigs have a long-standing reputation, the number of purebred Lingao pigs has dropped sharply in recent years due to the impact of foreign local pig breeds on the Hainan pig market and factors such as African swine fever. Research on the characteristic genetic resources of Lingao pigs and promoting the protection of purebred Lingao pigs are of great significance to the characteristic development of my country's pig breeding industry and the maintenance of the diversity of germplasm resource banks. In the long-term breeding process, unique genetic markers have been formed in the genome of the Lingao pig population, and these genetic markers can be used to accurately identify purebred Lingao pigs.

[0004] Single nucleotide polymorphism (SNP) is one of the most common forms of genetic variation. It is widely present in the genome of organisms and is also the most common genetic marker in the genome. Genome wide association study (GWAS) and selection signal analysis are effective methods for identifying specific SNP sites in biological populations. The combination of the two can further achieve the precise identification of specific SNP sites formed during the selection and breeding process of biological populations. The precise identification of purebred Lingao pigs at the genetic level using the unique SNP markers of the Lingao pig population is of great significance for the precise protection of the genetic resources of Hainan Lingao pigs. Summary of the invention

[0005] In order to overcome the deficiencies and shortcomings of the prior art, the primary purpose of the present invention is to provide a specific SNP site combination for identifying the Lingao pig breed.

[0006] Another object of the present invention is to provide an application of the above-mentioned specific SNP site combination.

[0007] Another object of the present invention is to provide a method for quickly identifying the Lingao pig breed.

[0008] The purpose of the present invention is achieved through the following technical solutions:

[0009] A specific SNP site combination for identifying the Lingao pig breed includes 61 SNP sites, named SNP sites 1-61, the physical positions of the 61 SNP sites are determined based on sequence comparison of the reference genome Sscrofa11.1 of the pig, and the position information and variation information of the 61 SNP sites are as follows:

[0010] SNP1 is located at 64741370bp on chromosome 1, and its mutation type is A / G;

[0011] SNP2 is located at 64771316bp on chromosome 1, and its mutation type is G / A;

[0012] SNP3 is located at 139752887bp on chromosome 1, and its mutation type is C / G;

[0013] SNP4 is located at 169195974bp on chromosome 1, and its mutation type is C / A;

[0014] SNP5 is located at 169215989bp on chromosome 1, and its mutation type is A / G;

[0015] SNP6 is located at 174613413bp on chromosome 1, and its mutation type is G / C;

[0016] SNP7 is located at 174623001bp on chromosome 1, and its mutation type is T / A;

[0017] SNP8 is located at 177743150bp on chromosome 1, and its mutation type is A / G;

[0018] SNP9 is located at 177836119bp on chromosome 1, and its mutation type is A / C;

[0019] SNP10 is located at 177836752bp on chromosome 1, and its mutation type is T / C;

[0020] SNP11 is located at 177839657bp on chromosome 1, and its mutation type is C / T;

[0021] SNP12 is located at 177937653bp on chromosome 1, and its mutation type is A / G;

[0022] SNP13 is located at 180517146bp on chromosome 1, and its mutation type is T / C;

[0023] SNP14 is located at 195820383bp on chromosome 1, and its mutation type is A / G;

[0024] SNP15 is located at 232406920bp on chromosome 1, and its mutation type is A / G;

[0025] SNP16 is located at 239555243bp on chromosome 1, and its mutation type is A / G;

[0026] SNP17 is located at 239558549bp on chromosome 1, and its mutation type is G / A;

[0027] SNP18 is located at 251247152bp on chromosome 1, and its mutation type is A / G;

[0028] SNP19 is located at 251431877bp on chromosome 1, and its mutation type is A / T;

[0029] SNP20 is located at 52507435bp on chromosome 3, and its mutation type is A / G;

[0030] SNP21 is located at 58390334bp on chromosome 3, and its mutation type is G / A;

[0031] SNP22 is located at 67522705bp on chromosome 3, and its mutation type is A / C;

[0032] SNP23 is located at 74217990bp on chromosome 3, and its mutation type is C / G;

[0033] SNP24 is located at 74236906bp on chromosome 3, and its mutation type is A / G;

[0034] SNP25 is located at 74250167bp on chromosome 3, and its mutation type is G / T;

[0035] SNP26 is located at 97670783bp on chromosome 3, and its mutation type is T / C;

[0036] SNP27 is located at 6273646bp on chromosome 5, and its mutation type is T / C;

[0037] SNP28 is located at 6274307bp on chromosome 5, and its mutation type is G / A;

[0038] SNP29 is located at 27586453bp on chromosome 6, and its mutation type is A / G;

[0039] SNP30 is located at 27588641bp on chromosome 6, and its mutation type is T / G;

[0040] SNP31 is located at 43834831bp on chromosome 6, and its mutation type is A / G;

[0041] SNP32 is located at 49174531bp on chromosome 6, and its mutation type is A / G;

[0042] SNP33 is located at 52193534bp on chromosome 6, and its mutation type is A / C;

[0043] SNP34 is located at 52261602bp on chromosome 6, and its mutation type is G / C;

[0044] SNP35 is located at 60450201bp on chromosome 6, and its mutation type is A / G;

[0045] SNP36 is located at 60476486bp on chromosome 6, and its mutation type is A / G;

[0046] SNP37 is located at 73469548bp on chromosome 6, and its mutation type is T / C;

[0047] SNP38 is located at 73700901bp on chromosome 6, and its mutation type is G / T;

[0048] SNP39 is located at 91187771bp on chromosome 6, and its mutation type is C / T;

[0049] SNP40 is located at 96273650bp on chromosome 6, and its mutation type is A / G;

[0050] SNP41 is located at 117205220bp on chromosome 6, and its mutation type is C / A;

[0051] SNP42 is located at 117208990bp on chromosome 6, and its mutation type is A / G;

[0052] SNP43 is located at 41997361bp on chromosome 8, and its mutation type is A / T;

[0053] SNP44 is located at 77471304bp on chromosome 8, and its mutation type is C / A;

[0054] SNP45 is located at 37767893bp on chromosome 12, and its mutation type is T / C;

[0055] SNP46 is located at 12560079bp on chromosome 13, and its mutation type is T / C;

[0056] SNP47 is located at 12590411bp on chromosome 13, and its mutation type is C / T;

[0057] SNP48 is located at 41667743bp on chromosome 13, and its mutation type is A / T;

[0058] SNP49 is located at 41713399bp on chromosome 13, and its mutation type is T / C;

[0059] SNP50 is located at 160260817bp on chromosome 13, and its mutation type is C / T;

[0060] SNP51 is located at 160263953bp on chromosome 13, and its mutation type is T / C;

[0061] SNP52 is located at 87138547bp on chromosome 14, and its mutation type is A / G;

[0062] SNP53 is located at 87149779bp on chromosome 14, and its mutation type is G / A;

[0063] SNP54 is located at 7519609bp on chromosome 17, and its mutation type is C / T;

[0064] SNP55 is located at 14497486bp on chromosome 17, and its mutation type is T / C;

[0065] SNP56 is located at 22299687bp on chromosome 17, and its mutation type is A / G;

[0066] SNP57 is located at 22305057bp on chromosome 17, and its mutation type is T / A;

[0067] SNP58 is located at 3820648bp on chromosome 18, and its mutation type is A / G;

[0068] SNP59 is located at 46306767bp on chromosome 18, and its mutation type is C / T;

[0069] SNP60 is located at 46309848bp on chromosome 18, and its mutation type is C / T;

[0070] SNP61 is located at 53236225bp on chromosome 18, and its mutation type is T / C.

[0071] Application of the specific SNP locus combination in identifying Lingao pig breeds;

[0072] Application of a product for detecting the above-mentioned specific SNP locus combination in identifying the Lingao pig breed;

[0073] The product is a reagent, a kit or a gene chip, etc.;

[0074] The reagents include at least one of primers, probes, etc.;

[0075] A molecular probe combination for identifying the Lingao pig breed, the molecular probe combination targets the above-mentioned specific SNP site combination and is used to detect the above-mentioned specific SNP site combination;

[0076] A gene chip for identifying the Lingao pig breed, the chip being loaded with the above molecular probe combination;

[0077] A method for quickly identifying the breed of Lingao pigs preferably comprises the following steps:

[0078] (1) Collecting tissue samples from the pigs to be tested and extracting DNA to obtain DNA samples from the pigs to be tested;

[0079] (2) performing gene sequencing on the DNA sample of the pig to be tested obtained in step (1) to obtain genotype data of the pig to be tested;

[0080] (3) The genotype data of the pig to be tested obtained in step (2) are combined with the genotype data of the known Lingao pig group and the control group, and the genotype data of the above-mentioned specific SNP site combination (61 SNP sites) are extracted. The combined and extracted genotype data are subjected to principal component analysis, and the principal component analysis graph is visualized by the R language ggplot2 package to determine whether the pig to be tested is a Lingao pig;

[0081] The DNA sample described in step (2) must first undergo quality inspection, and then undergo gene sequencing after passing the quality inspection;

[0082] The known Lingao pig group described in step (3) is a group consisting of multiple (≥15) purebred Lingao pigs, which may be the group consisting of the Lingao pigs in the embodiment, or a group consisting of Lingao pigs selected by the user, or a group of Lingao pigs in a public database;

[0083] The control group described in step (3) is a group of multiple pigs (≥5 pigs for a single breed group, ≥30 pigs for the entire group) other than Lingao pigs, which can be the control group group in the embodiment, or other non-Lingao pig groups selected by oneself, or non-Lingao pig groups in a public database;

[0084] The control group is preferably at least one of Ding'an pig, Tunchang pig, Wuzhishan pig, Bama Xiang pig, Diannan small-eared pig, Erhualian pig, Meishan pig, Anqing pig, Min pig, Tibetan Xiang pig, Landrace pig, Large White pig, Duroc pig and Asian wild boar;

[0085] The extraction and merging described in step (3) are preferably performed using Plink software;

[0086] The judgment method described in step (3) is:

[0087] When the results of principal component analysis show that the genetic distance between the pig to be tested and the known Lingao pig population is close, that is, they are clustered in one cluster, the pig to be tested is a Lingao pig; otherwise, it is a pig of other breeds.

[0088] Compared with the prior art, the present invention has the following advantages and effects:

[0089] (1) In order to achieve rapid and accurate identification of purebred Lingao pigs during the breeding process of Lingao pigs, the present invention, based on resequencing data, obtained a specific SNP site combination for identifying the Lingao pig breed by combining whole genome association analysis with selection signal analysis. This method further improves the accuracy of identifying the specific SNP site combination. The selected specific SNP site combination is more reliable in the identification of the Lingao pig breed.

[0090] (2) The SNP locus combination provided by the present invention has strong specificity and can be used for the scientific and accurate identification and genetic relationship analysis of purebred Lingao pigs, which is conducive to promoting the protection and development and utilization of Hainan's characteristic local pig varieties.

[0091] (3) The present invention provides a method for rapidly identifying purebred Lingao pigs. The method uses genomics and bioinformatics techniques to accurately identify purebred Lingao pigs, providing an effective scientific basis for the protection of the Lingao pig breed at the genetic level. BRIEF DESCRIPTION OF THE DRAWINGS

[0092] Figure 1 It is a Manhattan plot of the GWAS analysis results of the Lingao pig of the present invention.

[0093] Figure 2 It is a Manhattan plot of the selection signal analysis results of the Lingao pig of the present invention.

[0094] Figure 3 It is a distribution diagram of each specific SNP site disclosed in the present invention in each chromosome.

[0095] Figure 4 It is a genotype comparison diagram of different groups at each specific SNP site disclosed in the present invention.

[0096] Figure 5 It is a verification result diagram of the Lingao pig-specific SNP site combination disclosed in the present invention and the principal component analysis of other breeds. DETAILED DESCRIPTION

[0097] The present invention is further described in detail below in conjunction with embodiments and drawings, but the embodiments of the present invention are not limited thereto.

[0098] Example 1 Screening of Lingao Pig-Specific SNP Sites

[0099] 1. Genotype data acquisition

[0100] (1) The experimental population included 809 pigs of 15 breeds, including 11 local Chinese breeds, including four Hainan local breeds: Lingao pig (n=27, LG), Ding'an pig (n=24, DA), Tunchang pig (n=29, TC), and Wuzhishan pig (n=26, WZS); two South China breeds: Bama Xiang pig (n=6, BMX) and Diannan small-eared pig (n=33, DNXE); two Jianghai pig breeds: Erhualian pig (n=24, EHL) and Meishan pig (n=43, MS); one East China breed: Anqing pig (n=18, AQ); one North China breed: Min pig (n=18, MIN); and one plateau breed: Tibetan Xiang pig (n=118, ZX). There are three Western pig breeds, including Landrace (n=52, LR), Large White (n=177, LW), and Duroc (n=172, DU), and one wild boar breed, the Asian wild boar (n=42, AWB).

[0101] Table 1 Statistical table of experimental population

[0102] Serial number variety type Quantity (head) 1 Lingao Pig Hainan local pig 27 2 Dingan Pig Hainan local pig 24 3 Tunchang Pig Hainan local pig 29 4 Wuzhishan Pig Hainan local pig 26 5 Bama Xiang pig South China Pig 6 6 Yunnan Small-eared Pig South China Pig 33 7 Two-faced pig Jianghai Pig 24 8 Meishan Pig Jianghai Pig 43 9 Anqing Pig East China Pig 18 10 Minzhu North China Pig 18 11 Tibetan Pig Plateau Pig 118 12 Landrace pig Western breed pigs 52 13 Large White Pig Western breed pigs 177 14 Duroc pig Western breed pigs 172 15 Asian wild boar Wild Boar 42

[0103] (2) The genotype data of the Lingao pig, Ding'an pig, Tunchang pig, and Wuzhishan pig populations were directly obtained by resequencing. The specific method is as follows:

[0104] Ear tissue samples were collected from Lingao pigs, Ding'an pigs, Tunchang pigs, and Wuzhishan pigs, and the total DNA of the samples was extracted using the Omega tissue DNA extraction kit. The integrity and purity of the DNA of each sample were inspected by agarose gel electrophoresis and UV spectrophotometer. The DNA sample light absorption ratio (A260 / 280) was required to be between 1.8 and 2.0, and the concentration was ≥50 ng / μL. The DNA samples that passed the quality inspection were genotyped using the DNBSEQ sequencing platform (Shenzhen BGI Genomics Service Co., Ltd.) to obtain whole-genome genotype data. The sequencing depth was 10×, and a total of 40,201,194 SNP sites were obtained for 106 samples.

[0105] (3) The genotype data of other pigs were obtained from the PGIDB database.

[0106] 2. Genotype data processing

[0107] (1) The genotype data of each group were merged according to the conventional method using Plink software (command: --bmerge), and the merged genetic data were quality controlled. SNPs with a site detection rate lower than 95% (command: --geno0.05) and individuals with an individual detection rate lower than 90% (command: --mind 0.1) were removed. A total of 12,633,058 SNP sites shared by 809 individuals were obtained.

[0108] (2) A secondary quality control was then performed. Based on the original quality control conditions, loci with a minimum allele frequency of less than 1% and sex chromosome loci were removed (command: --maf 0.1 --chr 1-18). Finally, 12,631,911 high-quality SNP loci from 809 individuals were used for subsequent analysis.

[0109] 3. Screening of specific loci for Lingao pigs by genome-wide association analysis

[0110] Based on step 2, GEMMA software was used to conduct genome-wide association analysis based on a mixed linear model. The specific method was as follows: Lingao pigs were set as the experimental group, and other breeds were set as the control group for association analysis. The significant site threshold was set to 1e-25.

[0111] like Figure 1 As shown, the significant sites above the threshold line were selected as Group A of Lingao pig breed-specific sites.

[0112] 4. Select signal analysis to screen specific sites of Lingao pigs

[0113] Based on step 2, Plink software was used to convert the quality-controlled genotype data into vcf format (command: --recode vcf-iid), and VCFtools software was used to calculate the genetic differentiation index (Fst). The specific method was as follows: Lingao pigs were used as the experimental group, and other breeds were uniformly set as the control group. The sliding window size was set to 50,000 bp (command: --fst-window-size 50,000), and the sliding step was set to 10,000 bp (command: --fst-window-step 10,000), and the Fst values ​​of the analysis results were sorted from large to small.

[0114] like Figure 2 As shown, the sites in the top 1% window with the largest Fst value are group B of the Lingao pig breed-specific SNP sites.

[0115] 5. Screening of common specific loci for whole genome association analysis and genetic differentiation index

[0116] The common SNP sites of group A and group B were retained as group C. The SNP sites of group C were sorted according to their position in the genome, and the SNP sites with a distance of less than 1Mb were regarded as a significant QTL region. The top 1-8 sites with the most significant P value (obtained from the whole genome association analysis in step 3) in each QTL region were retained as the specific SNP sites of Lingao pigs.

[0117] The distribution of each specific SNP site screened in the genome is as follows Figure 3 The specific site information is shown in Table 2. The genotypes of different populations at each specific SNP site are shown in Figure 4 As shown in the figure, the rows represent different specific SNP sites, the columns represent all individuals used for analysis, and different breeds are represented by different colors above each column. The higher the frequency of the major allele of each SNP site, the closer it is to red. Figure 4 The results showed that the genotypes of the specific SNP loci of Lingao pigs were significantly different from those of other populations.

[0118] Table 2 Combination of 61 specific SNP molecular markers of Lingao pig

[0119] Serial number chromosome Physical Location Mutation type 1 1 64741370 A / G 2 1 64771316 G / A 3 1 139752887 C / G 4 1 169195974 C / A 5 1 169215989 A / G 6 1 174613413 G / C 7 1 174623001 T / A 8 1 177743150 A / G 9 1 177836119 A / C 10 1 177836752 T / C 11 1 177839657 C / T 12 1 177937653 A / G 13 1 180517146 T / C 14 1 195820383 A / G 15 1 232406920 A / G 16 1 239555243 A / G 17 1 239558549 G / A 18 1 251247152 A / G 19 1 251431877 A / T 20 3 52507435 A / G 21 3 58390334 G / A 22 3 67522705 A / C 23 3 74217990 C / G 24 3 74236906 A / G 25 3 74250167 G / T 26 3 97670783 T / C 27 5 6273646 T / C 28 5 6274307 G / A 29 6 27586453 A / G 30 6 27588641 T / G 31 6 43834831 A / G 32 6 49174531 A / G 33 6 52193534 A / C 34 6 52261602 G / C 35 6 60450201 A / G 36 6 60476486 A / G 37 6 73469548 T / C 38 6 73700901 G / T 39 6 91187771 C / T 40 6 96273650 A / G 41 6 117205220 C / A 42 6 117208990 A / G 43 8 41997361 A / T 44 8 77471304 C / A 45 12 37767893 T / C 46 13 12560079 T / C 47 13 12590411 C / T 48 13 41667743 A / T 49 13 41713399 T / C 50 13 160260817 C / T 51 13 160263953 T / C 52 14 87138547 A / G 53 14 87149779 G / A 54 17 7519609 C / T 55 17 14497486 T / C 56 17 22299687 A / G 57 17 22305057 T / A 58 18 3820648 A / G 59 18 46306767 C / T 60 18 46309848 C / T 61 18 53236225 T / C

[0120] Example 2 PCA visualization analysis of specific SNP loci combination identification of Tunlingao pigs

[0121] 1. Processing of genotype data

[0122] Referring to Example 1, the genotype data of 15 pig breeds (data of 809 individuals merged in step 2) in Example 1 were extracted using Plink software according to Table 2 to obtain genotype data corresponding to 61 specific SNP sites (named: D).

[0123] 2. Variety identification application

[0124] (1) Principal component analysis (PCA)

[0125] Use Plink software to perform PCA analysis on the above genotype data D. The output result prefix is ​​named lgpca. The relevant code is as follows:

[0126] plink --bfile D --pca 3 --out lgpca

[0127] plink: used to start Plink;

[0128] --bfile D: used to read the genotype file D in binary format;

[0129] --pca 3: used to perform principal component analysis and obtain the first three columns of eigenvector values;

[0130] --out lgpca: outputs the principal component analysis results. In this embodiment, the file prefix is ​​named lgpca.

[0131] (2) Visualization

[0132] The output result (lgpca.eigenvec) of step (1) is sorted in the data format through Excel. Specifically, the first column is set as the breed (Breed), the second column is the eigenvector value 1 (PC1), and the third column is the eigenvector value 2 (PC2).

[0133] Among them, in the lgpca.eigenvec file, the second column is the sample id, the third column is the eigenvector value 1, and the fourth column is the eigenvector value 2. When sorting data in Excel, the variety can be determined according to the sample id, and finally sorted into drawing data with the first column as the variety (Breed), the second column as the eigenvector value 1 (PC1), and the third column as the eigenvector value 2 (PC2).

[0134] Read the sorted data (named pca in this example) into R language, and use the ggplot2 package to draw a PCA analysis chart based on the eigenvector values ​​(PC1 and PC2) of the first two columns of principal component analysis. Set the X-axis to PC1 and the Y-axis to PC2, and use different colors to represent different breeds (Breed). The relevant code is as follows:

[0135] ggplot(pca,aes(x=PC1,y=PC2,color=Breed))+

[0136] geom_point(size=3)+

[0137] theme_bw() +

[0138] theme(panel.border=element_blank(), panel.grid.major=element_blank(), panel.grid.minor=element_blank(), axis.line=element_line(colour="black"))

[0139] ggplot(pca, aes(x=PC1, y=PC2, color=Breed)):

[0140] ggplot(): The function of the ggplot2 library is used to create graphic objects;

[0141] pca: object / data frame defining the data, containing the result data of principal component analysis;

[0142] aes(x=PC1, y=PC2, color=Breed): Aesthetic mapping function aes(), specifies how to map data to graphic attributes; x=PC1: maps the PC1 column in the data to the x-axis; y=PC2: maps the PC2 column in the data to the y-axis; color=Breed: uses the Breed column in the data to determine the color of the scatter points.

[0143] geom_point(size=3):

[0144] geom_point(): This is a function that adds a scatter point layer to the graph; size=3: sets the size of the scatter points to 3.

[0145] theme_bw():

[0146] Apply a theme, theme_bw() represents the "black and white" theme, which is a simple theme with a white background and grid lines;

[0147] theme(panel.border=element_blank(), panel.grid.major=element_blank(), panel.grid.minor=element_blank(), axis.line=element_line(colour="black")):

[0148] theme(): used to set or adjust non-data elements of the graphic; panel.border=element_blank(): removes the border of the graphic panel; panel.grid.major=element_blank(): removes major grid lines; panel.grid.minor=element_blank(): removes minor grid lines; axis.line=element_line(colour="black"): sets the axis line to black.

[0149] The results are as follows Figure 5 As shown, the 27 Lingao pigs clustered into one cluster, showing a close genetic distance, and a long genetic distance from the other 14 pig breeds, indicating that the Lingao purebred pigs can be accurately identified by combining 61 specific SNP loci with PCA analysis. Therefore, the 61 specific SNP loci provided by the present invention can be used to identify the breed of Lingao pigs.

[0150] Example 3 Application of Lingao pig-specific SNP locus combination in identification of purebred Lingao pigs

[0151] 1. Genotype data acquisition

[0152] Collect tissue samples from the pigs to be tested, extract DNA from the tissue samples, and obtain DNA samples from the pigs to be tested.

[0153] 2. Obtaining SNP data of the pigs to be tested

[0154] The DNA samples of the pigs to be tested are quality inspected, and after passing the quality inspection, they are sent to a gene sequencing company for gene sequencing to obtain the genotype data of the pigs to be tested.

[0155] 3. Processing of genotype data

[0156] Referring to Example 1, the genotype data of the pigs to be tested obtained in step 2 were merged with the genotype data of the known Lingao group (Table 1) and the control group (non-Lingao group in Table 1) through Plink software, and combined with Table 2, the genotype data corresponding to 61 specific SNP sites were extracted.

[0157] 4. Variety identification application

[0158] (1) Principal component analysis (PCA)

[0159] The specific method is the same as in Example 2;

[0160] (2) Visualization

[0161] The specific method is the same as Example 2.

[0162] (3) Result determination

[0163] When the principal component analysis results show that the genetic distance between the pig to be tested and the known Lingao pig population (Lingao pig population in Table 1) is close, that is, they are clustered in one cluster, the pig to be tested is Lingao pig; otherwise, it is other breeds of pig.

[0164] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be equivalent replacement methods and are included in the protection scope of the present invention.

Claims

1. Application of a product for detecting a specific SNP locus combination in identifying Lingao pig breeds, characterized in that: The specific SNP locus combination consists of 61 SNP loci, and the physical positions of the 61 SNP loci are based on the reference genome of the pig Sscrofa11.1 The position information and variation information of the 61 SNP sites are determined by sequence comparison as follows: SNP1 is located at 64741370bp on chromosome 1, and its mutation type is A / G; SNP2 is located at bp 64771316 on chromosome 1, and its mutation type is G / A; SNP3 is located at 139752887bp on chromosome 1, and its mutation type is C / G; SNP4 is located at 169195974bp on chromosome 1, and its mutation type is C / A; SNP5 is located at 169215989bp on chromosome 1, and its mutation type is A / G; SNP6 is located at 174613413bp on chromosome 1, and its mutation type is G / C; SNP7 is located at 174623001bp on chromosome 1, and its mutation type is T / A; SNP8 is located at 177743150bp on chromosome 1, and its mutation type is A / G; SNP9 is located at 177836119bp on chromosome 1, and its mutation type is A / C; SNP10 is located at 177836752bp on chromosome 1, and its mutation type is T / C; SNP11 is located at 177839657bp on chromosome 1, and its mutation type is C / T; SNP12 is located at 177937653bp on chromosome 1, and its mutation type is A / G; SNP13 is located at 180517146bp on chromosome 1, and its mutation type is T / C; SNP14 is located at 195820383bp on chromosome 1, and its mutation type is A / G; SNP15 is located at 232406920bp on chromosome 1, and its mutation type is A / G; SNP16 is located at 239555243bp on chromosome 1, and its mutation type is A / G; SNP17 is located at bp 239558549 on chromosome 1, and its mutation type is G / A; SNP18 is located at bp 251247152 on chromosome 1, and its mutation type is A / G; SNP19 is located at bp 251431877 on chromosome 1, and its mutation type is A / T; SNP20 is located at 52507435bp on chromosome 3, and its mutation type is A / G; SNP21 is located at 58390334bp on chromosome 3, and its mutation type is G / A; SNP22 is located at 67522705bp on chromosome 3, and its mutation type is A / C; SNP23 is located at 74217990bp on chromosome 3, and its mutation type is C / G; SNP24 is located at 74236906bp on chromosome 3, and its mutation type is A / G; SNP25 is located at 74250167bp on chromosome 3, and its mutation type is G / T; SNP26 is located at 97670783bp on chromosome 3, and its mutation type is T / C; SNP27 is located at bp 6273646 on chromosome 5, and its mutation type is T / C; SNP28 is located at bp 6274307 on chromosome 5, and its mutation type is G / A; SNP29 is located at 27586453bp on chromosome 6, and its mutation type is A / G; SNP30 is located at 27588641bp on chromosome 6, and its mutation type is T / G; SNP31 is located at 43834831bp on chromosome 6, and its mutation type is A / G; SNP32 is located at 49174531bp on chromosome 6, and its mutation type is A / G; SNP33 is located at 52193534bp on chromosome 6, and its mutation type is A / C; SNP34 is located at 52261602bp on chromosome 6, and its mutation type is G / C; SNP35 is located at 60450201bp on chromosome 6, and its mutation type is A / G; SNP36 is located at bp 60476486 on chromosome 6, and its mutation type is A / G; SNP37 is located at 73469548bp on chromosome 6, and its mutation type is T / C; SNP38 is located at 73700901bp on chromosome 6, and its mutation type is G / T; SNP39 is located at 91187771bp on chromosome 6, and its mutation type is C / T; SNP40 is located at 96273650bp on chromosome 6, and its mutation type is A / G; SNP41 is located at 117205220bp on chromosome 6, and its mutation type is C / A; SNP42 is located at 117208990bp on chromosome 6, and its mutation type is A / G; SNP43 is located at 41997361bp on chromosome 8, and its mutation type is A / T; SNP44 is located at 77471304bp on chromosome 8, and its mutation type is C / A; SNP45 is located at bp 37767893 on chromosome 12, and its mutation type is T / C; SNP46 is located at bp 12560079 on chromosome 13, and its mutation type is T / C; SNP47 is located at 12590411bp on chromosome 13, and its mutation type is C / T; SNP48 is located at 41667743bp on chromosome 13, and its mutation type is A / T; SNP49 is located at 41713399bp on chromosome 13, and its mutation type is T / C; SNP50 is located at 160260817bp on chromosome 13, and its mutation type is C / T; SNP51 is located at 160263953bp on chromosome 13, and its mutation type is T / C; SNP52 is located at 87138547bp on chromosome 14, and its mutation type is A / G; SNP53 is located at bp 87149779 on chromosome 14, and its mutation type is G / A; SNP54 is located at 7519609bp on chromosome 17, and its mutation type is C / T; SNP55 is located at 14497486bp on chromosome 17, and its mutation type is T / C; SNP56 is located at bp 22299687 on chromosome 17, and its mutation type is A / G; SNP57 is located at bp 22305057 on chromosome 17, and its mutation type is T / A; SNP58 is located at bp 3820648 on chromosome 18, and its mutation type is A / G; SNP59 is located at 46306767bp on chromosome 18, and its mutation type is C / T; SNP60 is located at 46309848bp on chromosome 18, and its mutation type is C / T; SNP61 is located at 53236225bp on chromosome 18, and its mutation type is T / C.

2. The use according to claim 1, characterized in that: The product is a kit or a gene chip.

3. The use according to claim 1, characterized in that: The product described is a reagent; The reagents include at least one of primers and probes.

4. A method for rapid identification of Lingao pig breeds, characterized in that The following steps are included: Collecting tissue samples from the pigs to be tested, and extracting DNA to obtain DNA samples from the pigs to be tested; (2) performing gene sequencing on the DNA sample of the pig to be tested obtained in step (1) to obtain genotype data of the pig to be tested; (3) merging the genotype data of the pig to be tested obtained in step (2) with the genotype data of the known Lingao pig group and the control group, and extracting the genotype data of the specific SNP site combination described in claim 1, performing principal component analysis on the combined and extracted genotype data, and visualizing the principal component analysis graph using the R language ggplot2 package to determine whether the pig to be tested is a Lingao pig; The judgment method described in step (3) is: When the principal component analysis results show that the genetic distance between the pig to be tested and the known Lingao pig group is close, that is, they are clustered together, the pig to be tested is a Lingao pig; otherwise, it is a pig of other breeds; the known Lingao pig group is a group composed of multiple purebred Lingao pigs, and the control group is a group of other pigs that are not Lingao pigs.

5. The method for rapid identification of Lingao pig breeds according to claim 4, characterized in that: The control group is at least one of Ding'an pig, Tunchang pig, Wuzhishan pig, Bama Xiang pig, Diannan small-eared pig, Erhualian pig, Meishan pig, Anqing pig, Min pig, Tibetan Xiang pig, Landrace pig, Large White pig, Duroc pig and Asian wild boar.

6. The method for rapid identification of Lingao pig breeds according to claim 4, characterized in that: The extraction and merging described in step (3) were performed using Plink software.

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