Primer set, kit, gene chip and method for detecting kunming dog breed

Abstract

The application provides a primer set, a kit, a gene chip and a method for detecting Kunming dog breeds, wherein the method for detecting Kunming dog breeds comprises the following steps: collecting gene information of a to-be-detected dog; amplifying the gene information; detecting the amplified gene information by using a preset primer set to obtain a preset number of to-be-detected gene locus information sets; comparing the to-be-detected gene locus information sets with reference gene locus information sets; and determining Kunming dog breed information corresponding to the to-be-detected dog according to the comparison result. By using the method, a small amount of high-frequency gene mutation sites are used to identify Kunming dogs, the identification result is directly obtained, the identification experience of related identification personnel is not needed, the detection accuracy is high, the repeatability is high, and the identification time is short.

Description

Technical Field

[0001] This application relates to the field of biotechnology, and in particular to primer sets, reagent kits, gene chips, and methods for detecting the Kunming dog breed. Background Technology

[0002] The Kunming dog is an excellent working dog successfully bred in my country. It is of medium size, with a symmetrical appearance, dense skeletal structure, compact body, well-developed muscles, good working performance, stable genetics, strong adaptability, and strong disease resistance. It has strong proactive defense, high alertness, good obedience, and good concentration, making it suitable for high-intensity work and an excellent working dog.

[0003] One of the prerequisites for the selection and breeding of superior Kunming dogs is accurate breed identification. Conventional identification methods mainly rely on phenotypic traits, but this method is prone to errors due to the influence of growth characteristics. While genomic analysis can provide accurate identification, it is costly. Given the high demand for Kunming dogs as working dogs, it is necessary to establish a rapid breed identification technique for Kunming dogs. Summary of the Invention

[0004] In view of this, embodiments of this application provide primer sets, reagent kits, gene chips, and methods for detecting the Kunming dog breed, in order to solve the above-mentioned problems existing in the prior art.

[0005] According to a first aspect of the embodiments of this application, a primer set for detecting the Kunming dog breed is provided, the primer set comprising at least one primer pair: (1) Primer pair 1, used to detect site 118194236 on chromosome 1, includes: Upstream primer 5'-TAATGAACCATAAAACGTCCGCGTCATCCCTTCCA-3'; Downstream primer 5'-AGGTGTTATATAACGTCGTTTTAGGACACCCTAAA-3'; (2) The second primer pair, used to detect the 1st chromosome, locus 118949304, includes: Upstream primer 5'-ACTGTTACTGTTGTTGTGGTCATTATTGACGAGAG-3'; Downstream primer 5'-CCTCTCAACACTACGGAAGTTCCGTTGTTCAAGTA-3'; (3) The third primer pair is used to detect the 1st chromosome, locus 119193059, including: Upstream primer 5'-TATGGTTTAAAGAACGGTCTGTCCTCCAAACCTCA-3'; Downstream primer 5'-CGTAATTATTTTACCGTGAAACAAAAAGTGAAAGA-3'; (4) The fourth primer pair is used to detect the 85347257 locus on chromosome 3, including: Upstream primer 5'-GACGTAGTGTTGTCATAGAATCGCCAAGAGGTCTC-3'; Downstream primer 5'-ACCCACCCCCGTGTGATGGTCGGTAACGTGTGACT-3'; (5) Primer pair 5, used to detect site 22662296 on chromosome 4, includes: Upstream primer 5'-ACAGGACAGTTTTGAAATGGGAGAGGGAGAGGAAA-3'; Downstream primer 5'-GTCGGTGCTATGGGAGTTGACTTCCGGGTACATAA-3'; (6) Primer pair 6, used to detect site 25280282 on chromosome 5, includes: Upstream primer 5'-TATAAGAACCTATTATAGTATCTATTATATCGTTC-3'; Downstream primer 5'-CACGGGGTTTAAACTCATTTTATCTTTTGCCAACA-3'; (7) Primer pair 7, used to detect site 25280282 on chromosome 5, includes: Upstream primer 5'-GGTTTGACTAGTGACCCGTACAAGTCTCAAGTGGG-3'; Downstream primer 5'-AGTGTTCTACTCTAGGTCCCTTTTCAGTGGGTCGC-3'; (8) Primer pair 8, used to detect site 75692716 on chromosome 6, includes: Upstream primer 5'-AGATTAAGTCTTTTACGACCGGATCAAATATCCAC-3'; Downstream primer 5'-TGTTCTTGAAATTAAATTCCTTCGACGCCACCAGA-3'; (9) Primer pair 9, used to detect site 11812516 on chromosome 7, includes: Upstream primer 5'-TATGACCCGCCCAGTACTTATACACTTTATTTGAT-3'; Downstream primer 5'-CCGCCTAATATTTTTCTTTAATTAAAAGGAGAACC-3'; (10) Primer pair 10, used to detect site 19242447 on chromosome 7, includes: Upstream primer 5'-CTGTAAAGAATATTTGTAAAATTCAGAAAAAGACT-3'; Downstream primer 5'-AAGTTAAAAACAGTAATAAAGAGATGGGAAGAGGT-3'; (11) Primer pair 11, used to detect site 33371309 on chromosome 10, includes: Upstream primer 5'-TCATCATTTGGTTTGTTTACGTTGATTGAGAAGGG-3'; Downstream primer 5'-GTGCATCGTATTCTATAATCGACCATCCTCGACTA-3'; (12) Primer pair 12, used to detect site 66455860 on chromosome 10, includes: Upstream primer 5'-TATCCGTGGTGACACTCTTAAAAAAAAAAATTAGG-3'; Downstream primer 5'-GTTAAGTCTCCGGACACTGTAAAGGAAGGAACCCT-3'; (13) Primer pair 13, used to detect site 38129850 on chromosome 17, includes: Upstream primer 5'-TTATCTACGAACGAACTAACTCTTTCTATTATGTA-3'; Downstream primer 5'-CAGTTATCTACTTTCTTCGGTCTCTATTCCGTGTT-3'; (14) Primer pair 14, used to detect site 52798015 on chromosome 17, includes: Upstream primer 5'-AACCCTAAAACTTCTATACTCCTGAAGTGACACTG-3'; Downstream primer 5'-CTAAGTTTCTAGTTTCACACCTTTTAATAGAGACC-3'; (15) Primer pair 15, used to detect chromosome 19, locus 37603608, includes: Upstream primer 5'-ACCCTTCCCAACTCCTCATCTTATCCACCATTTGC-3'; Downstream primer 5'-GGGTTTCCAAACAGTACCTTTAGGAGACCGAATAT-3'; (16) Primer pair 16, used to detect chromosome 19, locus 37776953, includes: Upstream primer 5'-ACTACCTTTTGTAACCACCCTTATACACCTAGGAT-3'; Downstream primer 5'-CCAATGAGATCTTAATGATCTACCATTCATCGATT-3'; (17) Primer pair 17, used to detect site 27382292 on chromosome 21, includes: Upstream primer 5'-AGGGTCGTGTCAGAACTCCTAGTCCTACATTCTAT-3'; Downstream primer 5'-TCGTTACTCGTATTTCAGATGGTATTCTGTTTCAC-3'; (18) Primer pair 18, used to detect chromosome 21, locus 30191215, includes: Upstream primer 5'-ATTTCTATGAAAGAGTCGGACATGAACGTGGTTGG-3'; Downstream primer 5'-CAGATACTATCCAGGGGGTCTTAAATAATGAAGTA-3'; (19) Primer pair 19, used to detect chromosome 21, locus 50334969, includes: Upstream primer 5'-TGACGCACTTCAACCGGCACTACACGAGTACCAGC-3'; Downstream primer 5'-CCCCCCACATCAGCCGTTGAGGACCTTTGTTCGTC-3'; (20) Primer pair 20, used to detect chromosome 23, locus 28512855, includes: Upstream primer 5'-GACTTTTGGAGGGTTGTGTCGGATACCAGACGGTA-3'; Downstream primer 5'-AACAGTAAGGGTCGATCCTGTAAGACCTGAAAGAT-3'; (21) Primer pair 21, used to detect site 9397982 on chromosome 24, includes: Upstream primer 5'-TCCGACCAGATTTAATCTTAGGTCCGTAACGACAC-3'; Downstream primer 5'-GGACATAAAGGTAGTGTGTGTCAGGGGTGTACACT-3'; (22) Primer pair 22, used to detect chromosome 24, locus 30600448, includes: Upstream primer 5'-GTGAATTTCTGTACAAGCTTGTGAACCATTCGGGG-3'; Downstream primer 5'-ACGAAAAATAAAACGTCACCATTCGTTTATTCCCT-3'; (23) Primer pair 23, used to detect site 6032411 on chromosome 27, includes: Upstream primer 5'-CATACCTCTTTTAGTCGAGGTCTCCACTCGGACTC-3'; Downstream primer 5'-AACCGTACTTTATCGCTTCTTTTCGTCTAGGTATC-3'; (24) Primer pair 24, used to detect chromosome 32, locus 2872926, includes: Upstream primer 5'-TGTTCCTCGAGACGGTCTCTCACGCGTTGTCCGAC-3'; Downstream primer 5'-CGGTGTATAATGAAGAAAGGAAAGGGTAAACCTAT-3'; (25) Primer pair 25, used to detect chromosome 32, locus 23029589, includes: Upstream primer 5'-TAACTGATCTGAACCGAAATAACCCTTAGAACTAT-3'; Downstream primer 5'-GACTGTGTTTCTTTCCGGAATAGAACCCAAGTCCT-3'; (26) Primer pair 26, used to detect chromosome 32, locus 24092241, includes: Upstream primer 5'-GTTGGTGAAGACGACTTTCTCCCTAACTCCAACAG-3'; Downstream primer 5'-ACCGGGGAGACTCTAGTCTTTTGGTGTGTCTGTTG-3'; (27) Primer pair 27, used to detect chromosome 32, locus 24169704, includes: Upstream primer 5'-TATTTAATATTACTCCCTTTCTTTTTACTGAATCT-3'; Downstream primer 5'-ACAGACGAATTAAGGGTTTCGATCTCTGTTATCCG-3'; (28) Primer pair 28, used to detect chromosome 33, locus 28219539, includes: Upstream primer 5'-GCAGACTACCGATCATCCGAGGAGAAATAATAAGT-3'; Downstream primer 5'-AAACCCCAGTGTGTACTGTGAAAAAAAAAAAACGG-3'; (29) Primer pair 29, used to detect site 9901059 on chromosome 37, includes: Upstream primer 5'-GGAGAAAACTAGAAAAAGTCTAAGGATCTCTACTA-3'; Downstream primer 5'-GAAGTTGTAAATGTGGAACCAGATAGGCACACAGA-3'; (30) Primer pair 30, used to detect chromosome 37, locus 18009157, includes: Upstream primer 5'-AAAACTTCCACCGTTTTTTGTTTTGGTTTTTGATT-3'; Downstream primer 5'-GGTAATACTTAGAAAATGTCTATTAGGTGGTTTAT-3'.

[0006] According to a second aspect of the embodiments of this application, a reagent kit for detecting the breed of Kunming dog is provided, comprising: The primer set provided in the first aspect of the embodiments of this application.

[0007] According to a third aspect of the embodiments of this application, a gene chip for detecting the Kunming dog breed is provided, comprising: At least two probes, wherein each pair of probes corresponds to a primer pair, and the primer pair is the primer pair in the primer set mentioned above.

[0008] According to a fourth aspect of the embodiments of this application, a method for detecting the breed of Kunming dogs is provided, comprising: Collect genetic information from the dogs to be tested; The gene information is amplified, and the amplified gene information is detected by a preset primer set to obtain a preset number of gene loci to be detected. The preset primer set includes the primer set provided in the first aspect of the embodiments of this application. The set of gene loci information to be detected is compared with the set of reference gene loci information; Based on the comparison results, the breed information of the Kunming dog corresponding to the dog to be tested was determined.

[0009] Optionally, collect the genetic information of the dog to be tested, including: Collect biological samples from the dogs to be tested; Genetic information of the dog to be tested was extracted from the biological sample.

[0010] Optionally, the set of gene loci information to be detected is compared with the set of reference gene loci information, including: The target gene locus information is determined from the set of gene locus information to be detected; Based on the target gene locus information to be detected, the target reference gene locus information is determined from the reference gene locus information set; Compare whether the target gene locus information to be detected and the target reference gene locus information are the same; If so, then the target gene locus information to be detected is confirmed to be successfully matched; If not, then the target gene locus information to be detected is determined to have failed to match.

[0011] Optionally, the breed information of the Kunming dog corresponding to the dog to be tested can be determined based on the comparison results, including: The number of gene loci that failed to match in the set of gene loci information to be detected is counted. If the number of gene loci to be detected is within the first detection interval, the dog to be detected is determined to be a purebred Kunming dog. If the number of gene loci to be detected is within the second detection interval, the dog to be detected is determined to be a hybrid Kunming dog; If the number of gene loci to be detected is within the third detection interval, it is determined that the dog to be detected is not a Kunming dog.

[0012] Optionally, the amplified gene information can be detected using a pre-set primer set, including: The amplified gene information is added to a kit or gene chip for detection, wherein the kit is the kit provided in the second aspect of the present application, and the gene chip is the gene chip provided in the third aspect of the present application.

[0013] According to a fifth aspect of the embodiments of this application, an apparatus for detecting the breed of Kunming dogs is provided, comprising: The acquisition module is configured to collect the genetic information of the dogs to be tested; The detection module is configured to amplify the gene information, detect the amplified gene information using a preset primer set, and obtain a preset number of gene locus information sets to be detected, wherein the preset primer set includes the primer set provided in the first aspect of the embodiments of this application. The comparison module is configured to compare the set of gene loci information to be detected with the set of reference gene loci information; The determination module is configured to determine the Kunming dog breed information corresponding to the dog to be tested based on the comparison results.

[0014] According to a sixth aspect of the present application, a computing device is provided, including a memory, a processor, and computer instructions stored in the memory and executable on the processor, wherein the processor executes the computer instructions to implement the steps of the method for detecting the breed of Kunming dogs.

[0015] According to a seventh aspect of the present application, a computer-readable storage medium is provided that stores computer instructions, which, when executed by a processor, implement the steps of the method for detecting the breed of Kunming dogs.

[0016] The method for detecting the breed of Kunming dog provided in this application involves collecting the genetic information of the dog to be tested; amplifying the genetic information; detecting the amplified genetic information using a preset primer set to obtain a preset number of gene loci information sets to be tested; comparing the gene loci information sets to be tested with a reference gene loci information set; and determining the Kunming dog breed information corresponding to the dog to be tested based on the comparison results.

[0017] One embodiment of this application realizes a method for identifying Kunming dogs using a small number of high-frequency gene mutation sites, which directly obtains identification results without relying on the identification experience of relevant identification personnel. The method has high detection accuracy, high repeatability, and short identification time. Attached Figure Description

[0018] Figure 1 This is a schematic diagram of upstream and downstream primer positioning target site information provided in an embodiment of this application; Figure 2 This is a flowchart of a method for detecting the breed of Kunming dogs provided in an embodiment of this application; Figure 3 This is a flowchart illustrating a method for detecting the breed of Kunming dogs according to an embodiment of this application; Figure 4 This is a schematic diagram of a device for detecting the breed of Kunming dogs according to an embodiment of this application; Figure 5 This is a structural block diagram of a computing device provided in one embodiment of this application. Detailed Implementation

[0019] Many specific details are set forth in the following description to provide a full understanding of this application. However, this application can be implemented in many other ways different from those described herein, and those skilled in the art can make similar extensions without departing from the spirit of this application; therefore, this application is not limited to the specific embodiments disclosed below.

[0020] The terminology used in one or more embodiments of this application is for the purpose of describing particular embodiments only and is not intended to limit the scope of one or more embodiments of this application. The singular forms “a,” “the,” and “the” used in one or more embodiments of this application and in the appended claims are also intended to include the plural forms unless the context clearly indicates otherwise. It should also be understood that the term “and / or” used in one or more embodiments of this application refers to and includes any or all possible combinations of one or more associated listed items.

[0021] It should be understood that although the terms first, second, etc., may be used to describe various information in one or more embodiments of this application, such information should not be limited to these terms. These terms are only used to distinguish information of the same type from one another. For example, first may also be referred to as second without departing from the scope of one or more embodiments of this application, and similarly, second may also be referred to as first. Depending on the context, the word "if" as used herein may be interpreted as "when," "when," or "in response to a determination."

[0022] The Kunming dog is a local breed, mainly produced in Yunnan. Through long-term selection and breeding by technicians, it has gradually developed into a unique working dog breed adapted to local environmental conditions. Due to its strong adaptability, high reproductive rate, and ease of training, it is one of the excellent genetic resources for working dogs.

[0023] One of the prerequisites for the selection and breeding of superior Kunming dogs is the accurate identification of breeding dogs. The current conventional identification method mainly combines epimorphological identification, but this method is prone to erroneous identification results due to the influence of growth characteristics. While identification using genomic analysis can be accurate, the identification cost is high.

[0024] Based on this, this application provides a primer set, reagent kit, gene chip and method for detecting the Kunming dog breed, a computing device, and a computer-readable storage medium, which will be described in detail in the following embodiments.

[0025] It should be noted that the canine reference genome used in this application embodiment is CanFam3.1, and the site information is a certain coordinate of a certain genome in the reference genome, such as sequence number 1, the site information is "1:118194236", that is, located on the first chromosome, at the 118194236th base.

[0026] Example 1 This embodiment provides a primer set for detecting the Kunming dog breed, which includes at least one of the following 30 primer pairs, as shown in Table 1 below: Table 1

[0027] In Table 1, the primer pair numbers represent the sequence information corresponding to each primer pair. The locus information indicates which genotype of DNA site the primer pair is used to detect. In the locus information, the number before the colon indicates which chromosome the locus is located on, and the number after the colon indicates the location of the locus on that chromosome. Taking primer pair 1 as an example, the locus information corresponding to this primer pair is "1:118194236". Here, the 1 before the colon indicates that the locus is on chromosome 1, and the 118194236 after the colon indicates that the locus is located at position 118194236 on chromosome 1.

[0028] Each primer pair includes an upstream primer and a downstream primer. A primer, at the initiation of polymerization, stimulates the synthesis of another macromolecule and is a sequence covalently linked to the reactant. In nucleic acid chemistry, a primer is a short single-stranded RNA (ribonucleic acid) or DNA (deoxyribonucleic acid) fragment that binds to a complementary region on the nucleic acid chain. Its function is to serve as the starting point for nucleotide polymerization. Nucleic acid polymerases can synthesize new nucleic acid chains starting from its 3' end. Artificially synthesized primers are widely used in polymerase chain reactions, sequencing, and probe synthesis.

[0029] The two ends of a DNA molecule are not the same. DNA replication always proceeds from the 5' end to the 3' end because DNA polymerase can only add nucleotides to the 3' end. So, whichever replicates first is upstream, and the one that replicates later is downstream. Upstream and downstream are relative concepts. The upstream primer is closer to the 5' end, and the downstream primer is closer to the 3' end.

[0030] Upstream and downstream primers are upstream and downstream gene fragments from the gene locus to be located. Their function is to locate and extract genotypic information from the corresponding loci. See also... Figure 1 , Figure 1 This is a schematic diagram of upstream and downstream primers for locating target site information provided in the embodiments of this application. When it is desired to extract the genotype of the target site information, the upstream and downstream gene fragments of the target site information can be located on the gene fragment through the upstream and downstream primers of the target site information, thereby extracting the genotype information of the target site information.

[0031] After extensive experimentation, 30 relatively stable gene loci associated with the Kunming dog were identified. These loci are: “1:118194236”, “1:118949304”, “1:119193059”, “3:85347257”, “4:22662296”, “5:25280282”, “5:33586501”, “6:75692716”, “7:11812516”, “7:19242447”, “10:33371309”, “10:66455860”, “17:38129850”, and “17:527980”. 15”, 19:37603608”, 19:37776953”, 21:27382292”, 21:30191215”, 21:50334969”, 23:28512855”, 24:9397982”, 24:30600448”, 27:6032411”, 32:2872926”, 32:23029589”, 32:24092241”, 32:24169704”, 33:28219539”, 37:9901059”, 37:18009157”.

[0032] The genotypes corresponding to the aforementioned 30 gene loci are relatively stable and not prone to mutation in a large amount of experimental data. Therefore, the breed information of the Kunming dog can be determined by using the genotype information corresponding to these 30 gene loci. For each of the 30 gene loci, this application provides upstream and downstream primers. Each primer pair corresponds to one locus. In practical applications, each primer pair can extract the genotype information of the corresponding locus. The primer set provided in this application for detecting the Kunming dog breed includes at least one of the aforementioned 30 primer pairs. Preferably, the primer set includes 30 primer pairs. These primer pairs can extract the genotype information from the 30 gene loci for subsequent breed detection.

[0033] In a preferred embodiment provided in this application, any 15 primer pairs from Table 1 above are used in the primer set for detecting the Kunming dog breed. The detection accuracy is 99.7% when the 15 primer pairs are used to detect the dog.

[0034] In another preferred embodiment provided in this application, any 20 primer pairs in Table 1 above are used in the primer set for detecting the Kunming dog breed. The detection accuracy is 99.95% when the 20 primer pairs are used to detect the dog.

[0035] In another preferred embodiment provided in this application, any 25 primer pairs in Table 1 above are used in the primer set for detecting the Kunming dog breed. The detection accuracy is 99.993% when the 25 primer pairs are used to detect the dog.

[0036] In another preferred embodiment provided in this application, the primer set for detecting the Kunming dog breed uses 30 primer pairs from Table 1 above. The detection accuracy is 99.999% when the 30 primer pairs are used to detect the dogs.

[0037] The primer set for detecting Kunming dogs provided in this application includes primer pairs corresponding to 30 gene loci. The primer set includes at least one of these 30 primer pairs. The primer set is used to extract genotype information of the corresponding gene loci, which facilitates the subsequent detection of genotype information of the target gene loci of the dogs to be tested. By designing different primer pairs for different gene loci, the corresponding gene loci can be located more accurately, the genotype information corresponding to the gene loci can be extracted, the extraction accuracy can be improved, and thus the accuracy of subsequent detection can be improved.

[0038] Example 2 One embodiment of this application also provides a kit for detecting the breed of Kunming dogs, wherein the kit includes the primer set for detecting the breed of Kunming dogs mentioned in the above embodiment. Furthermore, the more primer pairs in the primer set, the higher the accuracy for detecting the breed of Kunming dogs.

[0039] The kit provided in this application embodiment can be a PCR kit containing a primer set for detecting the Kunming dog breed. For example, in addition to the primer set mentioned above, the kit may also include test tubes or other suitable containers, reaction buffer, deoxyribonucleic acid, dideoxynucleotide triphosphate, polymerase and reverse enzyme, Dnase, RnAse inhibitor, DEPC-water and sterilizing agent, etc.

[0040] The kit for detecting the Kunming dog breed provided in this application includes at least one of 30 primer pairs, preferably 30 primer pairs. By placing the gene fragment of the dog to be tested into the kit, amplifying it in the kit, and then locating the corresponding gene locus through each primer pair, the genotype information of each gene locus is obtained. This information is then compared with the reference information of the Kunming dog breed to determine whether the dog to be tested is a Kunming dog.

[0041] Example 3 One embodiment of this application also provides a gene chip for detecting the Kunming dog breed.

[0042] Gene chips, also known as DNA chips, gene microarrays, oligonucleotide chips, or DNA microarrays, use hybridization sequencing, a method that determines nucleic acid sequences by hybridizing a set of nucleic acid probes with known sequences.

[0043] Specifically, a probe with a known target nucleotide sequence is fixed on the surface of a substrate. When the nucleic acid sequence in the solution is complementary to the nucleic acid probe at the corresponding position on the gene chip, a set of probe sequences that are completely complementary are obtained by means of the probe position. Based on this, the sequence of the target nucleic acid can be reconstructed.

[0044] Gene chips immobilize a large number of probe molecules on a support, which then hybridize with labeled sample molecules. By detecting the hybridization signal intensity of each probe molecule, the quantity and sequence information of the sample molecules can be obtained. In simpler terms, through microfabrication, tens of thousands, or even millions, of DNA fragments with specific sequences (gene probes) are systematically arranged and immobilized on supports such as silicon wafers or glass slides to form a two-dimensional DNA probe array. Gene chips are primarily used for gene detection.

[0045] The gene chip for detecting Kunming dogs provided in this application includes multiple probes, each with a primer pair. These primer pairs correspond to the primer pairs in Example 1. For example, a gene chip may have 30 probes, each corresponding to a gene locus. Each probe also has upstream and downstream primers corresponding to that gene locus. Once the gene information of the dog to be tested is captured by the probes, the upstream and downstream primers on the probes match the upstream and downstream gene fragments of the corresponding gene locus in the gene information, thereby determining the corresponding gene locus. Then, the genotype information of the corresponding gene locus is extracted, thus achieving the purpose of extracting genotype information corresponding to the locus information.

[0046] In practical applications, the number of probes on a gene chip can be tens of thousands, hundreds of thousands, or even millions. In this application, the specifications of the gene chip are not limited. It is only required that the probes of the gene chip include at least one of the 30 upstream and downstream primer pairs used to detect the Kunming dog breed. It should be noted that in order to improve the detection accuracy of the Kunming dog breed, the more of the 30 primer pairs used to detect the Kunming dog breed deployed on the gene chip, the better. Preferably, 30 primer pairs used to detect the Kunming dog breed are deployed.

[0047] In the embodiments provided in this application, a silicon crystal gene chip is provided, with a size of 0.5 cm * 0.5 cm, which contains a total of 250 * 250 = 62,500 loci. Each locus includes a pair of probes. Thirty loci are selected from the 62,500 loci for detecting the Kunming dog breed, which correspond one-to-one with the loci in Table 1 above. One of the probes in each locus contains the upstream primer corresponding to the locus, and the other contains the downstream primer corresponding to the locus. When a gene fragment is dropped onto the gene chip, the pair of probes at each locus can capture the corresponding gene locus, thereby extracting the genotype information corresponding to the gene locus.

[0048] The gene chip for detecting Kunming dog breed provided in this application embodiment is deployed at multiple sites, each site including a pair of probes. Each pair of probes corresponds to upstream and downstream primers for the same gene site. By using a pair of probes at each site, the corresponding gene position in the DNA can be captured and located, thereby extracting the genotype information of that gene position. This facilitates subsequent comparison with reference information of Kunming dog breed, thereby determining whether the dog to be tested is a Kunming dog.

[0049] Example 4 This application also provides a method for detecting the breed of Kunming dogs. See [link to relevant documentation]. Figure 2 , Figure 2 The flowchart illustrates a method for detecting the breed of Kunming dogs according to an embodiment of this application, specifically including the following steps: Step 202: Collect the genetic information of the dog to be tested.

[0050] Specifically, the dog to be tested refers to a dog that needs to be determined to be a Kunming dog. In the method provided in this application, the breed information of the dog to be tested is detected.

[0051] Genetic information specifically refers to the DNA (deoxyribonucleic acid) information of the dog to be tested. In practical applications, the DNA information of each dog is unique. By collecting the DNA information of the dog to be tested and then comparing and analyzing the DNA information, the breed information of the dog to be tested can be determined.

[0052] Specifically, the genetic information of the dogs to be tested is collected, including: Collect biological samples from the dogs to be tested; Genetic information of the dog to be tested was extracted from the biological sample.

[0053] In practical applications, the DNA information of the dog to be tested is contained in its biological sample. A biological sample can be understood as a complete or partial remnant of a living animal or plant, such as blood, bodily fluids, hair, etc., that carries DNA information. After obtaining the biological sample, the genetic information of the dog to be tested can be extracted from it.

[0054] In one specific embodiment provided in this application, taking blood as an example of biological sample, whole blood is extracted from the dog to be tested. During sample collection, to ensure DNA stability, heparin sodium anticoagulant is added to the collected whole blood, and the blood collection tube is stored in a liquid nitrogen container. Upon returning to the laboratory, the DNA preservation solution is transferred to a specific sample storage tube at a ratio of 1:3 (whole blood to DNA preservation solution) and stored at -80°C. Then, phenol is used... chloroform Isoamyl alcohol extraction method is used to extract DNA from whole blood to obtain the genetic information of the dogs to be tested.

[0055] In another specific embodiment provided in this application, taking the oral swab of the dog to be tested as an example, the oral swab of the dog to be tested is collected and placed into the test tube to be tested. The collected oral swab is centrifuged or compressed to make the sample meet the experimental requirements (at least 1 ml).

[0056] Nucleic acid extraction was then performed on the samples using a dual-plate extraction method. The difference was that the elution buffer in the EB plate was 70 μL of ultrapure water, facilitating subsequent concentration treatment if the sample concentration was low. The extracted samples were temporarily stored at 4°C until the samples were removed from the machine. In practical applications, for quantitative determination of DNA concentration, if the concentration was greater than 17.2 ng / μL, subsequent experiments could proceed. If the concentration was between 17.2 ng / μL and 6 ng / μL, concentration was considered before proceeding to subsequent experiments. If the concentration was less than 6 ng / μL, resampling was required.

[0057] Step 204: Amplify the gene information, and detect the amplified gene information using a preset primer set to obtain a preset number of gene loci to be detected.

[0058] The preset primer set includes the primer set mentioned in Example 1.

[0059] The above steps have extracted gene information. At this point, the number of gene information is relatively small. In order to ensure that each primer pair can match the gene information, the gene information needs to be amplified. For example, if there are 10 gene information in the extracted sample solution, after amplification, the concentration of gene information in the sample solution is increased. Then, the sample solution after amplification is used for detection to obtain a preset number of gene loci information sets to be detected.

[0060] The gene locus information to be detected specifically refers to the gene locus information used to detect the breed information of Kunming dogs. This gene locus information to be detected is the actual gene locus information of the dog to be detected. The preset number is the number of gene loci used in this application to detect the breed information of Kunming dogs. If 15 gene loci are used to detect the breed information of Kunming dogs, the preset number is 15; if 30 gene loci are used to detect the breed information of Kunming dogs, the preset number is 30.

[0061] In practical applications, amplified gene information can be detected using a pre-set primer set. The vector for the pre-set primer set can be a reagent kit or a gene chip. Specifically, the detection of amplified gene information using a pre-set primer set includes: The amplified gene information is added to a kit or gene chip for detection, wherein the kit is the one mentioned in Example 2, and the gene chip is the one mentioned in Example 3.

[0062] In practical applications, gene information needs to be detected based on a preset primer set. The preset primer set is the primer set of one of the above embodiments. The number of primer pairs in the primer set can be set according to the actual situation. Preferably, in order to improve the detection accuracy, 30 primer pairs are selected.

[0063] The preset primer set is set in the reagent kit or gene chip. The reagent kit or gene chip used in this embodiment is the reagent kit in Embodiment 2 above or the gene chip in Embodiment 3 above. For relevant information about the reagent kit or gene chip, please refer to the above description, which will not be repeated here.

[0064] The method provided in this application allows for the acquisition of a predetermined set of gene loci information to be detected. This set of gene loci information is used for genotype data comparison in subsequent testing processes.

[0065] Step 206: Compare the set of gene loci information to be detected with the set of reference gene loci information.

[0066] The reference gene locus information set specifically refers to the comparison data of Kunming dogs obtained after a large number of experiments, as shown in Table 2: Table 2

[0067] Referring to Table 2 above, taking the locus information "1:118194236" as an example, the genotype information corresponding to this gene locus for a purebred Kunming dog is "G / G"; taking the locus information "24:9397982" as an example, the genotype information corresponding to this gene locus for a purebred Kunming dog is "C / C"...

[0068] Once the set of gene loci information for the dog to be tested is obtained, the set of gene loci information is compared with the reference gene loci information to determine the breed information of the dog to be tested.

[0069] The comparison of the set of gene loci information to be detected with the set of reference gene loci information includes: The target gene locus information is determined from the set of gene locus information to be detected; Based on the target gene locus information to be detected, the target reference gene locus information is determined from the reference gene locus information set; Compare whether the target gene locus information to be detected and the target reference gene locus information are the same; If so, then the target gene locus information to be detected is confirmed to be successfully matched; If not, then the target gene locus information to be detected is determined to have failed to match.

[0070] In practical applications, each gene locus in the set of gene locus information to be detected needs to be compared with its corresponding reference gene locus information. Taking locus information "1:118194236" as an example, the target gene locus information for the dog to be detected corresponding to this locus information is "C / C", while the target reference gene locus information corresponding to this locus information is "G / G". Since the target gene locus information and the target reference gene locus information are different, the target gene locus information to be detected fails to match. Taking locus information "7:19242447" as an example, the target gene locus information for the dog to be detected corresponding to this locus information is "T / T", while the target reference gene locus information corresponding to this locus information is "T / T". Since the target gene locus information and the target reference gene locus information are the same, the target gene locus information to be detected matches successfully.

[0071] Step 208: Determine the Kunming dog breed information corresponding to the dog to be tested based on the comparison results.

[0072] After the above comparison process, the breed information of the Kunming dog corresponding to the dog to be tested can be determined based on the comparison results. Specifically, the breed information of the Kunming dog includes three types: purebred Kunming dog, mixed-breed Kunming dog, and non-Kunming dog.

[0073] In practical applications, the breed information of the Kunming dog corresponding to the dog to be tested is determined based on the comparison results, including: The number of gene loci that failed to match in the set of gene loci information to be detected is counted. If the number of gene loci to be detected is within the first detection interval, the dog to be detected is determined to be a purebred Kunming dog. If the number of gene loci to be detected is within the second detection interval, the dog to be detected is determined to be a hybrid Kunming dog; If the number of gene loci to be detected is within the third detection interval, it is determined that the dog to be detected is not a Kunming dog.

[0074] In the method provided in this application, the breed information of the dog to be tested, namely the Kunming dog, is determined by determining the number of gene sites that failed to match. The first detection interval, the second detection interval, and the third detection interval specifically refer to the range of the number of gene sites that failed to match. In practical applications, the first detection interval can be 0-2, the second detection interval can be 3-4, and the third detection interval can be 5 or more.

[0075] When the number of failed gene loci to be tested is within the first detection interval, it means that the number of failed gene loci to be tested is 0, 1 or 2. At this time, it means that the dog to be tested is a purebred Kunming dog, that is, there are a maximum of 2 different genotypes among the gene loci to be tested.

[0076] When the number of failed gene loci to be tested is within the second detection interval, it means that there are 3 or 4 failed gene loci to be tested. At this time, the dog to be tested can still be identified as a Kunming dog, but the dog to be tested is a hybrid Kunming dog.

[0077] When the number of failed gene loci to be tested is in the third detection interval, it means that the number of failed gene loci to be tested is greater than or equal to 5. At this time, the dog to be tested can be identified as a non-Kunming dog.

[0078] The method for detecting the breed of Kunming dog provided in this application involves collecting the genetic information of the dog to be tested; amplifying the genetic information; detecting the amplified genetic information using a preset primer set to obtain a preset number of gene loci to be tested, wherein the preset primer set includes the primer set in Example 1; comparing the set of gene loci to be tested with a reference gene loci information set; and determining the Kunming dog breed information corresponding to the dog to be tested based on the comparison results. This method solves the problems of high cost and long time required for whole-genome sequencing of Kunming dogs. By identifying Kunming dogs using a small number of high-frequency gene loci, the identification results are obtained directly, without relying on the identification experience of the personnel, avoiding human factors, and achieving high accuracy, high repeatability, and short identification time.

[0079] The following is in conjunction with the appendix Figure 3 Taking the method for detecting the breed of Kunming dogs provided in this application as an example, the method for detecting the breed of Kunming dogs will be further explained. Among other things, Figure 3The present application provides a flowchart of a method for detecting the breed of Kunming dogs according to an embodiment of this application, which specifically includes the following steps: Step 302: Collect biological samples from the dog to be tested.

[0080] Step 304: Extract the genetic information of the dog to be tested from the biological sample.

[0081] Step 306: Amplify the gene information to obtain amplified gene information.

[0082] Step 308: Add the amplified gene information to the gene chip for detection to obtain a preset set of gene loci information to be detected.

[0083] The gene chip includes 30 probe pairs, each corresponding to an upstream and downstream primer for a single site, for a total of 30 primer pairs. Specific information about the 30 primer pairs can be found in Table 1 above, and will not be repeated here.

[0084] Step 310: Obtain the reference gene locus information set for Kunming dogs.

[0085] Step 312: Compare the set of gene loci to be detected with the set of reference gene loci, and count the number of gene loci that failed to match.

[0086] Step 314: Determine the breed information of the dog to be tested based on the number of unmatched gene loci.

[0087] Corresponding to the above method embodiments, this application also provides an embodiment of a device for detecting the breed of Kunming dogs. Figure 4 A schematic diagram of a device for detecting the breed of Kunming dogs, according to an embodiment of this application, is shown. Figure 4 As shown, the device includes: The acquisition module 402 is configured to acquire the genetic information of the dog to be tested; The detection module 404 is configured to amplify the gene information, detect the amplified gene information using a preset primer set, and obtain a preset number of gene locus information sets to be detected, wherein the preset primer set includes the primer set in the above embodiment 1. The comparison module 406 is configured to compare the set of gene loci information to be detected with the set of reference gene loci information; The determination module 408 is configured to determine the Kunming dog breed information corresponding to the dog to be detected based on the comparison results.

[0088] Optionally, the acquisition module 402 is further configured to: Collect biological samples from the dogs to be tested; Genetic information of the dog to be tested was extracted from the biological sample.

[0089] Optionally, the comparison module 406 is further configured to: The target gene locus information is determined from the set of gene locus information to be detected; Based on the target gene locus information to be detected, the target reference gene locus information is determined from the reference gene locus information set; Compare whether the target gene locus information to be detected and the target reference gene locus information are the same; If so, then the target gene locus information to be detected is confirmed to be successfully matched; If not, then the target gene locus information to be detected is determined to have failed to match.

[0090] Optionally, the determining module 408 is further configured to: The number of gene loci that failed to match in the set of gene loci information to be detected is counted. If the number of gene loci to be detected is within the first detection interval, the dog to be detected is determined to be a purebred Kunming dog. If the number of gene loci to be detected is within the second detection interval, the dog to be detected is determined to be a hybrid Kunming dog; If the number of gene loci to be detected is within the third detection interval, it is determined that the dog to be detected is not a Kunming dog.

[0091] Optionally, the detection module 404 is further configured to: The amplified gene information is added to a kit or gene chip for detection, wherein the kit is the kit in Example 2 above, and the gene chip is the gene chip in Example 3 above.

[0092] The device for detecting the breed of Kunming dogs provided in this application collects the genetic information of the dog to be tested; amplifies the genetic information; detects the amplified genetic information using a preset primer set to obtain a preset number of gene loci to be tested, wherein the preset primer set includes the primer set in Embodiment 1; compares the set of gene loci to be tested with a reference gene loci information set; and determines the Kunming dog breed information corresponding to the dog to be tested based on the comparison results. This device solves the problems of high cost and long time required for whole-genome sequencing of Kunming dogs. By identifying Kunming dogs using a small number of high-frequency gene loci, it directly obtains identification results without relying on the identification experience of the personnel, avoiding human factors, and achieving high detection accuracy, high repeatability, and short identification time.

[0093] The above is a schematic diagram of a device for detecting the breed of Kunming dogs according to this embodiment. It should be noted that the technical solution of this device for detecting the breed of Kunming dogs belongs to the same concept as the technical solution of the method for detecting the breed of Kunming dogs described above. Details not described in detail in the technical solution of the device for detecting the breed of Kunming dogs can be found in the description of the technical solution of the method for detecting the breed of Kunming dogs described above.

[0094] Figure 5 A structural block diagram of a computing device 500 according to an embodiment of this application is shown. The components of the computing device 500 include, but are not limited to, a memory 510 and a processor 520. The processor 520 is connected to the memory 510 via a bus 530, and a database 550 is used to store data.

[0095] The computing device 500 also includes an access device 540, which enables the computing device 500 to communicate via one or more networks 560. Examples of these networks include a Public Switched Telephone Network (PSTN), a Local Area Network (LAN), a Wide Area Network (WAN), a Personal Area Network (PAN), or a combination of communication networks such as the Internet. The access device 540 may include one or more of any type of wired or wireless network interface (e.g., a Network Interface Card (NIC)), such as an IEEE 802.11 Wireless Local Area Network (WLAN) interface, a Wi-MAX interface, an Ethernet interface, a Universal Serial Bus (USB) interface, a cellular network interface, a Bluetooth interface, a Near Field Communication (NFC) interface, and so on.

[0096] In one embodiment of this application, the aforementioned components of the computing device 500 and Figure 5 Other components, not shown, can also be connected to each other, for example, via a bus. It should be understood that... Figure 5 The block diagram of the computing device shown is for illustrative purposes only and is not intended to limit the scope of this application. Those skilled in the art can add or replace other components as needed.

[0097] The computing device 500 can be any type of stationary or mobile computing device, including mobile computers or mobile computing devices (e.g., tablet computers, personal digital assistants, laptop computers, notebook computers, netbooks, etc.), mobile phones (e.g., smartphones), wearable computing devices (e.g., smartwatches, smart glasses, etc.) or other types of mobile devices, or stationary computing devices such as desktop computers or PCs. The computing device 500 can also be a mobile or stationary server.

[0098] The processor 520 executes the computer instructions to implement the steps of the method for detecting the breed of Kunming dogs.

[0099] The above is a schematic representation of a computing device according to this embodiment. It should be noted that the technical solution of this computing device and the technical solution of the method for detecting the breed of Kunming dogs described above belong to the same concept. Details not described in detail in the technical solution of the computing device can be found in the description of the technical solution of the method for detecting the breed of Kunming dogs described above.

[0100] An embodiment of this application also provides a computer-readable storage medium storing computer instructions that, when executed by a processor, implement the steps of the method for detecting the breed of Kunming dogs as described above.

[0101] The above is an illustrative embodiment of a computer-readable storage medium. It should be noted that the technical solution of this storage medium belongs to the same concept as the technical solution of the method for detecting the breed of Kunming dogs described above. Details not described in detail in the technical solution of the storage medium can be found in the description of the technical solution of the method for detecting the breed of Kunming dogs described above.

[0102] The foregoing has described specific embodiments of this application. Other embodiments are within the scope of the appended claims. In some cases, the actions or steps recited in the claims may be performed in a different order than that shown in the embodiments and may still achieve the desired results. Furthermore, the processes depicted in the drawings do not necessarily require the specific or sequential order shown to achieve the desired results. In some embodiments, multitasking and parallel processing are also possible or may be advantageous.

[0103] The computer instructions include computer program code, which may be in the form of source code, object code, executable file, or some intermediate form. The computer-readable medium may include: any entity or device capable of carrying the computer program code, recording media, USB flash drive, portable hard drive, magnetic disk, optical disk, computer memory, read-only memory (ROM), random access memory (RAM), electrical carrier signals, telecommunication signals, and software distribution media, etc. It should be noted that the content contained in the computer-readable medium may be appropriately added to or subtracted according to the requirements of legislation and patent practice in the jurisdiction. For example, in some jurisdictions, according to legislation and patent practice, computer-readable media may not include electrical carrier signals and telecommunication signals.

[0104] It should be noted that, for the sake of simplicity, the foregoing method embodiments are all described as a series of actions. However, those skilled in the art should understand that this application is not limited to the described order of actions, as some steps may be performed in other orders or simultaneously according to this application. Furthermore, those skilled in the art should also understand that the embodiments described in the specification are preferred embodiments, and the actions and modules involved are not necessarily essential to this application.

[0105] In the above embodiments, the descriptions of each embodiment have different focuses. For parts not described in detail in a certain embodiment, please refer to the relevant descriptions of other embodiments.

[0106] The preferred embodiments disclosed above are merely illustrative of this application. The optional embodiments do not exhaustively describe all details, nor do they limit the invention to the specific implementations described. Clearly, many modifications and variations can be made based on the content of this application. These embodiments are selected and specifically described in this application to better explain the principles and practical applications of this application, thereby enabling those skilled in the art to better understand and utilize this application. This application is limited only by the claims and their full scope and equivalents.

Claims

1. A primer set for detecting the Kunming dog breed, characterized in that, The primer set includes the following primer pairs: (1) Primer pair 1, used to detect site 118194236 on chromosome 1, includes: Upstream primer 5'-TAATGAACCATAAAACGTCCGCGTCATCCCTTCCA-3'; Downstream primer 5'-AGGTGTTATATAACGTCGTTTTAGGACACCCTAAA-3'; (2) The second primer pair, used to detect the 1st chromosome, locus 118949304, includes: Upstream primer 5'-ACTGTTACTGTTGTTGTGGTCATTATTGACGAGAG-3'; Downstream primer 5'-CCTCTCAACACTACGGAAGTTCCGTTGTTCAAGTA-3'; (3) The third primer pair is used to detect the 1st chromosome, locus 119193059, including: Upstream primer 5'-TATGGTTTAAAGAACGGTCTGTCCTCCAAACCTCA-3'; Downstream primer 5'-CGTAATTATTTTACCGTGAAACAAAAAGTGAAAGA-3'; (4) The fourth primer pair is used to detect the 85347257 locus on chromosome 3, including: Upstream primer 5'-GACGTAGTGTTGTCATAGAATCGCCAAGAGGTCTC-3'; Downstream primer 5'-ACCCACCCCCGTGTGATGGTCGGTAACGTGTGACT-3'; (5) Primer pair 5, used to detect site 22662296 on chromosome 4, includes: Upstream primer 5'-ACAGGACAGTTTTGAAATGGGAGAGGGAGAGGAAA-3'; Downstream primer 5'-GTCGGTGCTATGGGAGTTGACTTCCGGGTACATAA-3'; (6) Primer pair 6, used to detect site 25280282 on chromosome 5, includes: Upstream primer 5'-TATAAGAACCTATTATAGTATCTATTATATCGTTC-3'; Downstream primer 5'-CACGGGGTTTAAACTCATTTTATCTTTTGCCAACA-3'; (7) Primer pair 7, used to detect site 25280282 on chromosome 5, includes: Upstream primer 5'-GGTTTGACTAGTGACCCGTACAAGTCTCAAGTGGG-3'; Downstream primer 5'-AGTGTTCTACTCTAGGTCCCTTTTCAGTGGGTCGC-3'; (8) Primer pair 8, used to detect site 75692716 on chromosome 6, includes: Upstream primer 5'-AGATTAAGTCTTTTACGACCGGATCAAATATCCAC-3'; Downstream primer 5'-TGTTCTTGAAATTAAATTCCTTCGACGCCACCAGA-3'; (9) Primer pair 9, used to detect site 11812516 on chromosome 7, includes: Upstream primer 5'-TATGACCCGCCCAGTACTTATACACTTTATTTGAT-3'; Downstream primer 5'-CCGCCTAATATTTTTCTTTAATTAAAAGGAGAACC-3'; (10) Primer pair 10, used to detect site 19242447 on chromosome 7, includes: Upstream primer 5'-CTGTAAAGAATATTTGTAAAATTCAGAAAAAGACT-3'; Downstream primer 5'-AAGTTAAAAACAGTAATAAAGAGATGGGAAGAGGT-3'; (11) Primer pair 11, used to detect site 33371309 on chromosome 10, includes: Upstream primer 5'-TCATCATTTGGTTTGTTTACGTTGATTGAGAAGGG-3'; Downstream primer 5'-GTGCATCGTATTCTATAATCGACCATCCTCGACTA-3'; (12) Primer pair 12, used to detect site 66455860 on chromosome 10, includes: Upstream primer 5'-TATCCGTGGTGACACTCTTAAAAAAAAAAATTAGG-3'; Downstream primer 5'-GTTAAGTCTCCGGACACTGTAAAGGAAGGAACCCT-3'; (13) Primer pair 13, used to detect site 38129850 on chromosome 17, includes: Upstream primer 5'-TTATCTACGAACGAACTAACTCTTTCTATTATGTA-3'; Downstream primer 5'-CAGTTATCTACTTTCTTCGGTCTCTATTCCGTGTT-3'; (14) Primer pair 14, used to detect site 52798015 on chromosome 17, includes: Upstream primer 5'-AACCCTAAAACTTCTATACTCCTGAAGTGACACTG-3'; Downstream primer 5'-CTAAGTTTCTAGTTTCACACCTTTTAATAGAGACC-3'; (15) Primer pair 15, used to detect chromosome 19, locus 37603608, includes: Upstream primer 5'-ACCCTTCCCAACTCCTCATCTTATCCACCATTTGC-3'; Downstream primer 5'-GGGTTTCCAAACAGTACCTTTAGGAGACCGAATAT-3'; (16) Primer pair 16, used to detect chromosome 19, locus 37776953, includes: Upstream primer 5'-ACTACCTTTTGTAACCACCCTTATACACCTAGGAT-3'; Downstream primer 5'-CCAATGAGATCTTAATGATCTACCATTCATCGATT-3'; (17) Primer pair 17, used to detect site 27382292 on chromosome 21, includes: Upstream primer 5'-AGGGTCGTGTCAGAACTCCTAGTCCTACATTCTAT-3'; Downstream primer 5'-TCGTTACTCGTATTTCAGATGGTATTCTGTTTCAC-3'; (18) Primer pair 18, used to detect chromosome 21, locus 30191215, includes: Upstream primer 5'-ATTTCTATGAAAGAGTCGGACATGAACGTGGTTGG-3'; Downstream primer 5'-CAGATACTATCCAGGGGGTCTTAAATAATGAAGTA-3'; (19) Primer pair 19, used to detect chromosome 21, locus 50334969, includes: Upstream primer 5'-TGACGCACTTCAACCGGCACTACACGAGTACCAGC-3'; Downstream primer 5'-CCCCCCACATCAGCCGTTGAGGACCTTTGTTCGTC-3'; (20) Primer pair 20, used to detect chromosome 23, locus 28512855, includes: Upstream primer 5'-GACTTTTGGAGGGTTGTGTCGGATACCAGACGGTA-3'; Downstream primer 5'-AACAGTAAGGGTCGATCCTGTAAGACCTGAAAGAT-3'; (21) Primer pair 21, used to detect site 9397982 on chromosome 24, includes: Upstream primer 5'-TCCGACCAGATTTAATCTTAGGTCCGTAACGACAC-3'; Downstream primer 5'-GGACATAAAGGTAGTGTGTGTCAGGGGTGTACACT-3'; (22) Primer pair 22, used to detect chromosome 24, locus 30600448, includes: Upstream primer 5'-GTGAATTTCTGTACAAGCTTGTGAACCATTCGGGG-3'; Downstream primer 5'-ACGAAAAATAAAACGTCACCATTCGTTTATTCCCT-3'; (23) Primer pair 23, used to detect site 6032411 on chromosome 27, includes: Upstream primer 5'-CATACCTCTTTTAGTCGAGGTCTCCACTCGGACTC-3'; Downstream primer 5'-AACCGTACTTTATCGCTTCTTTTCGTCTAGGTATC-3'; (24) Primer pair 24, used to detect chromosome 32, locus 2872926, includes: Upstream primer 5'-TGTTCCTCGAGACGGTCTCTCACGCGTTGTCCGAC-3'; Downstream primer 5'-CGGTGTATAATGAAGAAAGGAAAGGGTAAACCTAT-3'; (25) Primer pair 25, used to detect chromosome 32, locus 23029589, includes: Upstream primer 5'-TAACTGATCTGAACCGAAATAACCCTTAGAACTAT-3'; Downstream primer 5'-GACTGTGTTTCTTTCCGGAATAGAACCCAAGTCCT-3'; (26) Primer pair 26, used to detect chromosome 32, locus 24092241, includes: Upstream primer 5'-GTTGGTGAAGACGACTTTCTCCCTAACTCCAACAG-3'; Downstream primer 5'-ACCGGGGAGACTCTAGTCTTTTGGTGTGTCTGTTG-3'; (27) Primer pair 27, used to detect chromosome 32, locus 24169704, includes: Upstream primer 5'-TATTTAATATTACTCCCTTTCTTTTTACTGAATCT-3'; Downstream primer 5'-ACAGACGAATTAAGGGTTTCGATCTCTGTTATCCG-3'; (28) Primer pair 28, used to detect chromosome 33, locus 28219539, includes: Upstream primer 5'-GCAGACTACCGATCATCCGAGGAGAAATAATAAGT-3'; Downstream primer 5'-AAACCCCAGTGTGTACTGTGAAAAAAAAAAAACGG-3'; (29) Primer pair 29, used to detect site 9901059 on chromosome 37, includes: Upstream primer 5'-GGAGAAAACTAGAAAAAGTCTAAGGATCTCTACTA-3'; Downstream primer 5'-GAAGTTGTAAATGTGGAACCAGATAGGCACACAGA-3'; (30) Primer pair 30, used to detect chromosome 37, locus 18009157, includes: Upstream primer 5'-AAAACTTCCACCGTTTTTTGTTTTGGTTTTTGATT-3'; Downstream primer 5'-GGTAATACTTAGAAAATGTCTATTAGGTGGTTTAT-3'.

2. A reagent kit for detecting the breed of Kunming dogs, characterized in that, Includes the primer set as described in claim 1.

3. A method for detecting the breed of Kunming dogs, characterized in that, include: Collect genetic information from the dogs to be tested; The gene information is amplified, and the amplified gene information is detected by a preset primer set to obtain a preset number of gene loci to be detected, wherein the preset primer set includes the primer set described in claim 1. The set of gene loci information to be detected is compared with the set of reference gene loci information; Based on the comparison results, the breed information of the Kunming dog corresponding to the dog to be tested was determined.

4. The method as described in claim 3, characterized in that, Collect the genetic information of the dogs to be tested, including: Collect biological samples from the dogs to be tested; Genetic information of the dog to be tested was extracted from the biological sample.

5. The method as described in claim 3, characterized in that, The set of gene loci information to be detected is compared with the set of reference gene loci information, including: The target gene locus information is determined from the set of gene locus information to be detected; Based on the target gene locus information to be detected, the target reference gene locus information is determined from the reference gene locus information set; Compare whether the target gene locus information to be detected and the target reference gene locus information are the same; If so, then the target gene locus information to be detected is confirmed to be successfully matched; If not, then the target gene locus information to be detected is determined to have failed to match.

6. The method as described in claim 5, characterized in that, Based on the comparison results, the breed information of the Kunming dog corresponding to the dog to be tested was determined, including: The number of gene loci that failed to match in the set of gene loci information to be detected is counted. If the number of gene loci to be detected is within the first detection interval, the dog to be detected is determined to be a purebred Kunming dog. If the number of gene loci to be detected is within the second detection interval, the dog to be detected is determined to be a hybrid Kunming dog; If the number of gene loci to be detected is within the third detection interval, it is determined that the dog to be detected is not a Kunming dog.

7. The method as described in claim 3, characterized in that, The amplified gene information was detected using a pre-set primer set, including: The amplified gene information is added to a kit or gene chip for detection, wherein the kit is the kit described in claim 2, and the gene chip is the gene chip described in claim 3.

8. A device for detecting the breed of Kunming dogs, characterized in that, include: The acquisition module is configured to collect the genetic information of the dogs to be tested; The detection module is configured to amplify the gene information, detect the amplified gene information using a preset primer set, and obtain a preset number of gene locus information sets to be detected, wherein the preset primer set includes the primer set described in claim 1. The comparison module is configured to compare the set of gene loci information to be detected with the set of reference gene loci information; The determination module is configured to determine the Kunming dog breed information corresponding to the dog to be tested based on the comparison results.

9. A computing device, comprising: Memory and processor; The memory is used to store computer-executable instructions, and the processor is used to execute the computer-executable instructions, which, when executed by the processor, implement the steps of the method according to any one of claims 3 to 7.

10. A computer-readable storage medium storing computer-executable instructions that, when executed by a processor, implement the steps of the method according to any one of claims 3 to 7.

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