Application of SNP (Single Nucleotide Polymorphism) marker related to horse body length and chest circumference characters

Abstract

The invention relates to an application of an SNP (Single Nucleotide Polymorphism) marker related to a horse body length and chest circumference character. The application specifically comprises the following applications: (1) identifying or assisting in identifying the horse body length or chest circumference character; (2) preparing a product for identifying or assisting in identifying the body length or chest circumference character of the horse; (3) breeding horses with slender figures and large chest circumference; (4) preparing and breeding products of horses with slender figures and large chest circumference; the SNP marker is located at the 76th bp, the 788th bp and the 230th bp of DNA of the No.15 chromosome of a horse genome, an allele is A or G, and rs ID is rs1136176196. By detecting the genotype of the SNP marker provided by the invention to assist in breeding of breeding horses, AA genotype individuals can be directionally selected and reserved in the foal stage, and the AA genotype individuals are used as racing horses or racing horse breeding horses for culture after a quick-step race of a 1000 / 5000 m race.

Description

Technical Field

[0001] The invention belongs to the field of molecular biology, and particularly relates to the application of SNP markers related to horse body length and chest girth traits. Background Art

[0002] China has one of the longest histories of horse breeding in the world. However, with social development and the widespread mechanization of the economy, the role of horses has gradually been replaced by machinery, and the horse industry has shifted from everyday uses in military and agricultural fields to recreational uses such as equestrianism and horse racing, as well as production for dairy and meat. The Xinjiang Uyghur Autonomous Region boasts the largest horse population in my country and boasts a rich equine resource, including four local breeds, such as the Kazakh horse, and two cultivated breeds, such as the Ili horse. In recent years, equestrian sports have rapidly developed in the Xinjiang Uyghur Autonomous Region, with a growing number of professional competitions. The horse industry has also gradually developed towards specialization, scale, and intensification. Currently, the majority of the sport horses employed by equestrian clubs across China come from the Xinjiang Uyghur Autonomous Region.

[0003] Bortala Mongol Autonomous Prefecture (Botala Prefecture) is a strategically located region on my country's western frontier, nestled in the heart of the Altai-Tianshan equine industry belt. Bordering traditional horse-breeding regions such as Altay, Tacheng, and Yining, Bortala Prefecture boasts a long history of horse breeding and a rich cultural heritage, providing a unique foundation for the development of its equine industry. Bortala Prefecture has long bred Kazakh horses, known for their strength and tolerance for roughage. However, their competitive abilities lag significantly behind those of leading domestic and international breeds, making them unable to meet the demands of modern competitive racing. In recent years, Bortala Prefecture has improved the Kazakh horse through the introduction of thoroughbreds and other leading breeds, including the Ili horse. The resulting hybrids have seen significant improvements in conformation, size, performance, and competitive performance.

[0004] A horse's body conformation significantly influences its quality and performance, and is a key factor in determining its competitive performance. Studies have shown that height, length, and chest circumference are significantly correlated with horse racing performance. For example, in the 1000-meter trot, higher height, longer length, and larger chest circumference, as well as smaller tube circumference, chest circumference ratio, and tube circumference ratio, are associated with shorter race times. In the 5000-meter trot, higher height, larger chest circumference, longer length, and smaller tube circumference ratio are associated with shorter race times. Scientific breeding and genetic improvement can cultivate horse breeds with superior conformation, not only improving overall performance but also better meeting market demand and increasing economic benefits. With the rapid development of molecular biology techniques, molecular genetic markers and marker-assisted selection have been increasingly widely studied and applied in livestock and poultry breeding, opening up new avenues for fundamentally improving equine genetic quality and breeding high-performance horses. Therefore, discovering SNP markers related to horse body size traits and applying them to the breeding of large-bodied, high-performance horses will significantly improve the efficiency and accuracy of breeding and accelerate the process of horse genetic improvement. Summary of the Invention

[0005] The present invention has screened for a single nucleotide polymorphism (SNP) marker significantly associated with body length and chest girth in horses. Detecting the genotype of this SNP marker can assist in the selection and breeding of racehorses or stallions, allowing targeted selection of individuals with the target genotype during the foal stage for breeding as reserve racehorses or stallions. Based on this, the present invention provides applications for SNP markers associated with body length and chest girth in horses.

[0006] In a first aspect, the present invention provides a use of a SNP marker associated with horse body length and chest girth traits in any one of (1) to (4): (1) Identify or assist in identifying the body length or chest girth of horses; (2) Preparation of products for identifying or assisting in identifying the body length or chest girth traits of horses; (3) Breeding horses with slender bodies and large chest circumference; (4) Preparation of products for breeding horses with slender bodies and large chest circumference; The SNP marker is located at 76,788,230 bp of chromosome 15 DNA of the horse genome, the allele is A or G, and the rsID is rs1136176196.

[0007] In a second aspect, the present invention provides a reagent for detecting SNP markers associated with horse body length and chest girth traits, as used in any one of (1) to (4): (1) Identify or assist in identifying the body length or chest girth of horses; (2) Preparation of products for identifying or assisting in identifying the body length or chest girth traits of horses; (3) Breeding horses with slender bodies and large chest circumference; (4) Preparation of products for breeding horses with slender bodies and large chest circumference; The SNP marker is located at 76,788,230 bp of chromosome 15 DNA of the horse genome, the allele is A or G, and the rsID is rs1136176196.

[0008] In a third aspect, the present invention provides a primer set for amplifying SNP markers associated with horse body length and chest girth traits, as used in any one of (1) to (4): (1) Identify or assist in identifying the body length or chest girth of horses; (2) Preparation of products for identifying or assisting in identifying the body length or chest girth traits of horses; (3) Breeding horses with slender bodies and large chest circumference; (4) Preparation of products for breeding horses with slender bodies and large chest circumference; The SNP marker is located at 76,788,230 bp of chromosome 15 DNA of the horse genome, the allele is A or G, and the rsID is rs1136176196.

[0009] In some embodiments of the present invention, the product includes one of a kit, a chip, and an instrument.

[0010] In a fourth aspect, the present invention provides a method for identifying or assisting in identifying the body length or chest girth traits of a horse, comprising: Detect the genotypes of SNP markers related to horse body length and chest girth traits in the genome of the test individuals; Predict the body length of the test individual when it grows up based on the genotype: If the genotype of the SNP marker associated with horse body length in the genome of the test individual is the AA genotype, it is predicted that the test individual will grow up to be a horse with a slender body and a large chest circumference; If the genotype of the SNP marker related to horse body length in the genome of the test individual is the AG genotype, it is predicted that the test individual will grow up to be a horse with a short body and a small chest circumference; The SNP marker is located at 76,788,230 bp of chromosome 15 DNA of the horse genome, the allele is A or G, and the rsID is rs1136176196.

[0011] In a fifth aspect, the present invention provides a method for breeding racehorses, comprising: Detect the genotypes of SNP markers related to horse body length and chest girth traits in the genome of the test individuals; If the genotype of the SNP marker related to horse body length in the genome of the test individual is AA genotype, the test individual will be bred and managed as a reserve racehorse; If the SNP marker related to horse body length in the genome of the test individual is AG genotype or GG genotype, the test individual will be eliminated; The SNP marker is located at 76,788,230 bp of chromosome 15 DNA of the horse genome, the allele is A or G, and the rsID is rs1136176196.

[0012] In a sixth aspect, the present invention provides a method for breeding racehorse stallions, comprising: Detect the genotypes of SNP markers related to horse body length and chest girth traits in the genome of the test individuals; If the genotype of the SNP marker related to horse body length in the genome of the test individual is AA genotype, the test individual will be bred and managed as a reserve racehorse; If the SNP marker related to horse body length in the genome of the test individual is AG or GG genotype, the test individual will be eliminated; The SNP marker is located at 76,788,230 bp of chromosome 15 DNA of the horse genome, the allele is A or G, and the rsID is rs1136176196.

[0013] In some embodiments of the present invention, the method for detecting the genotype of SNP markers related to horse body length and chest circumference traits in the genome of the test individual is any one of whole genome sequencing, PCR, gene chip, and SNaPshot.

[0014] In some embodiments of the present invention, the subject is a foal.

[0015] In some embodiments of the present invention, the test individual or its parent is a horse from the Bortala region.

[0016] Compared with the prior art, the present invention has at least the following beneficial effects: (1) The present invention screened a SNP marker associated with the body length trait of horses in Bortala Prefecture. The body length of horses with different genotypes of this SNP marker was significantly different; the average body length of individuals with the AA genotype was 145.4 cm, and the average body length of individuals with the AG genotype was 137.3 cm. The AA genotype was the dominant genotype, and the corresponding individuals had a slender body shape.

[0017] (2) This SNP marker can also be used to select for the chest girth trait of horses in the Bortala region. Horses with different genotypes of this SNP marker have significant differences in chest girth. The average chest girth of individuals with the AA genotype is 179.74 cm, and the average chest girth of individuals with the AG genotype is 172.83 cm. The AA genotype is the dominant genotype, and the corresponding individuals have both large chest girth traits.

[0018] (3) By detecting the genotype of the SNP marker provided by the present invention to assist in stallion breeding, the body length and chest circumference of the horse can be effectively increased by selecting stallions with the AA genotype in the foal stage, thereby realizing early selection of stallions, accelerating the genetic improvement progress and breeding process of stallions in the Bozhou area, and obtaining a specialized horse breed with a slender body and a large chest circumference.

[0019] (4) By detecting the genotype of the SNP marker provided by the present invention to assist stallion breeding, individuals with the AA genotype can be selectively selected at the foal stage to be trained as reserve racehorses for the 1000 / 5000 m race. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the following briefly introduces the drawings required for use in the description of the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without creative work.

[0021] Figure 1 :Selection pressure among populations on the chromosomal region where the rs1136176196 locus is located.

[0022] Figure 2 :Selection pressure on the chromosomal region where the rs1136176196 locus is located among extreme individuals. DETAILED DESCRIPTION

[0023] To make the objectives, technical solutions, and advantages of the present invention more clear, the technical solutions of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, but not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts shall fall within the scope of protection of the present invention.

[0024] The present invention screens out a SNP marker significantly associated with the body length and chest girth traits of horses. The SNP marker is located at 76,788,230 bp of chromosome 15 DNA of the horse genome, the allele is A or G, and the rs ID is rs1136176196.

[0025] As used herein, the term "genotype" refers to the allele combination of SNPs in the cells of an organism; "AA genotype" refers to the alleles of the SNP marker being A and A; the term "GG genotype" refers to the alleles of the SNP marker being G and G; and the term "AG genotype" refers to the alleles of the SNP marker being A and G.

[0026] As used herein, the term "chest girth" refers to the chest girth trait in livestock body size traits, measured as the length perpendicularly around the posterior edge of the shoulder blade. The terms "small chest girth" and "large chest girth" are used herein to describe the relative size of chest girth in adult horses across all genotypes of this linked SNP marker.

[0027] In this article, the term "body length" refers to the straight-line distance from the front edge of the shoulder to the back edge of the hip; the terms "slender body" and "short body" are used to describe the relative size of the body length of adult horses within the range of all genotype individuals of this linked SNP marker.

[0028] Herein, the terms "site", "SNP site" and "SNP marker" have the same meaning.

[0029] As used herein, the term "subject" is defined as the biological individual from which the test sample originates.

[0030] For the purposes of this article, the term "foal" is defined as an immature equine individual whose body size continues to develop and change until adulthood.

[0031] The present study found that the distribution of the three genotypes of this SNP marker varied significantly across horse breeds and individuals with extreme body lengths. The AA genotype was dominant in first-generation hybrids, advanced hybrids, and thoroughbreds, while the AG genotype was predominant in Kazakh horses. In Ili horses, the frequencies of the three genotypes were similar. In the extremely large group, the frequency of the AA genotype was 100%, significantly higher than the 57.1% in the extremely small group.

[0032] In horses in Bortala Prefecture, the AA genotype of this SNP marker corresponds to individuals with slender body and large chest circumference, with an average body length of 145.4 cm and chest circumference of 179.74 cm; the AG genotype corresponds to individuals with small body length and small chest circumference, with an average body length of 137.3 cm and chest circumference of 172.83 cm.

[0033] The present invention can determine the genotype of the SNP marker by directly detecting the SNP marker through whole genome sequencing or preparing a reagent for detecting the SNP marker, thereby realizing application in any of the following scenarios: (1) predicting the body length or chest girth traits of horses; (2) breeding racehorses with slender bodies and large chest girth at the foal stage, and then selectively selecting trotting racehorses for 1000 / 5000 m races; (3) breeding racehorse stallions at the foal stage for breeding horses with body length or chest girth traits, as well as improving horse breeds related to body length and chest girth traits, especially improving the germplasm resources of horses in Bortala area.

[0034] Based on this, the present invention provides a SNP marker associated with horse body length and chest girth traits, and its detection reagent and amplification primer set for use in any one of (1) to (4): (1) Identify or assist in identifying the body length or chest girth of horses; (2) Preparation of products for identifying or assisting in identifying the body length or chest girth traits of horses; (3) Breeding horses with slender bodies and large chest circumference; (4) Preparation of products for breeding horses with slender bodies and large chest circumference; The SNP marker is located at 76,788,230 bp of chromosome 15 DNA of the horse genome, the allele is A or G, and the rsID is rs1136176196.

[0035] In some embodiments of the present invention, the product includes one of a kit, a chip, and an instrument.

[0036] The present invention provides a method for identifying or assisting in identifying the body length or chest girth traits of a horse, comprising: Detect the genotypes of SNP markers related to horse body length and chest girth traits in the genome of the test individuals; Predict the body length of the test individuals when they grow up based on their genotype.

[0037] This method can be directly used for breeding decisions (e.g., screening reserve racehorses), for scientific research (analyzing the strength of associations between genotype and phenotype), or to assist veterinarians or breeders in developing personalized management plans (e.g., providing high-nutrient feed to foals predicted to be "slender"). For example, the test subject is a foal. If the genotype of the SNP marker associated with horse length in the test subject's genome is the AA genotype, the test subject is predicted to grow up to be a slender horse with a large chest girth. If the genotype of the SNP marker associated with horse length in the test subject's genome is the AG genotype, the test subject is predicted to grow up to be a short horse with a small chest girth.

[0038] The present invention provides a method for breeding racehorses, comprising: Detect the genotypes of SNP markers related to horse body length and chest girth traits in the genome of the test individuals; If the genotype of the SNP marker related to horse body length in the genome of the test individual is AA genotype, the test individual will be bred and managed as a reserve racehorse; If the SNP marker associated with horse body length in the genome of the test individual is AG or GG genotype, the test individual will be eliminated.

[0039] The body length trait of a foal is not yet established, making it difficult to predict its adult length. Selecting racehorses based on their phenotype after adulthood prevents them from reaching their full potential due to a lack of targeted training during their early years. Similarly, training all horses as racehorses during their early years and then eliminating unqualified ones after adulthood results in a significant waste of training resources. The present invention performs genotyping testing on foals under 24 months of age, allowing for the early elimination of inferior individuals, reducing subsequent rearing costs and increasing the proportion of high-quality racehorses.

[0040] In some embodiments, the test individuals are bred and managed as reserve racehorses, including: providing targeted training (such as endurance and speed training) and nutritional support (high-protein feed, trace element supplementation) to AA genotype foals; tracking and recording the body length and competition performance (such as speed and endurance indicators) of the foals after adulthood, and promptly eliminating unqualified ones.

[0041] The present invention also provides a method for breeding racehorse stallions, comprising: Detect the genotypes of SNP markers related to horse body length and chest girth traits in the genome of the test individuals; If the genotype of the SNP marker related to horse body length in the genome of the test individual is AA genotype, the test individual will be bred and managed as a reserve racehorse; If the SNP marker associated with horse body length in the genome of the test individual is AG or GG genotype, the test individual will be eliminated.

[0042] Phenotypic selection of stallions is typically performed when the horses are 3-4 years old, which prolongs the breeding cycle and reduces selection efficiency. Furthermore, body length and chest circumference phenotypes are susceptible to environmental factors such as rearing conditions and training level, and may not accurately reflect the stallion's genetic potential, causing selection results to deviate from actual genetic advantage. The present invention can determine the suitability of a test individual for breeding and management as a racehorse stallion by detecting the genotype of SNP markers within a few days of birth.

[0043] In some embodiments of the present invention, the method for detecting the genotype of SNP markers related to horse body length and chest circumference traits in the genome of the test individual is any one of whole genome sequencing, PCR, gene chip, and SNaPshot.

[0044] In some embodiments of the present invention, the subject is a foal. Preferably, the subject is no older than 24 months; more preferably, the subject is no older than 12 months; further, the subject is no older than 1 month.

[0045] In some embodiments of the present invention, the test individual or its parent is a horse from the Bortala region.

[0046] The technical solution of the present invention is described in detail below through specific embodiments: Example This study uses genomic population evolution and selection analysis to identify genetic markers associated with body size traits in local horse breeds. These markers can be used to improve the accuracy of body length selection in horses in Bortala Prefecture. The specific steps are as follows: 1. Materials and Methods 1.1 Blood collection and body measurement of horses in Bortala Prefecture Study subjects: 56 adult horses were randomly selected from Bortala Mongol Autonomous Prefecture, Xinjiang Uygur Autonomous Region, including 30 first-generation hybrid horses, 12 high-generation hybrid horses, and 14 Kazakh horses. First-generation hybrid horses were first-generation hybrids derived from crossing a bred horse (e.g., a Thoroughbred, Ili, or similar breed) with a Kazakh horse from Bortala Prefecture as the dam. High-generation hybrid horses were derived from crossing a first-generation hybrid horse with the aforementioned sires for multiple generations.

[0047] All blood samples were collected from the jugular vein, anticoagulated with EDTA, and stored at -20°C for 10× genome sequencing. The horses' height (vertical height from the top of the carapace to the ground), body length (straight line distance from the front edge of the shoulder to the back edge of the hip), chest circumference (vertical length around the back edge of the shoulder blade), and canal circumference (horizontally around the thinnest point of the upper third of the foreleg canal) were measured. The body length ratio, chest circumference ratio, and canal circumference ratio were calculated using the following formulas: Body length ratio (%) = body length / body height × 100%; Chest circumference ratio (%) = chest circumference / body height × 100%; Tube circumference rate (%) = tube circumference / body height × 100%.

[0048] 1.2 Blood total DNA extraction and quality control DNA was extracted using the Cowin CWE9600 fully automated nucleic acid extraction instrument and the accompanying Cowin CWE9600 Magbead Blood DNA Kit using magnetic beads. To the corresponding positions of a 96-well deep-well plate, 20 μL of Proteinase K, 300 μL of blood sample, 200 μL of Buffer mL, 750 μL of Buffer KL, 750 μL of Buffer CW1, 750 μL of Buffer GW2, 750 μL of Buffer MW3, and 100 μL of Buffer EB were added. The magnetic rod holder was inserted into the 96-well deep-well plate and the Cowin Blood 300 program was run. After approximately 23 minutes, 310 μL of the thoroughly mixed isopropanol and magnetic bead mixture was added according to the instrument prompt and the program was continued. The program concluded after approximately 35 minutes, and the eluted product was transferred to a 1.5 mL centrifuge tube for cryopreservation.

[0049] Take 2 μL of the eluted product and apply it to a 1.5% agarose gel. Perform electrophoresis at 150 V for 25 min. Place the gel in a UV analyzer. If only one band is observed, the DNA sample is considered to have good integrity. Use a NanoDrop 2000 Nucleic Acid Protein Analyzer to test the purity of the eluted product. If the A260 / A280 ratio is within the range of 1.7-2.1 and the A260 / A230 ratio is within the range of 1.8-2.2, the DNA sample purity meets the requirements for library construction.

[0050] 1.3 Library construction and sequencing Genomic DNA (>1 μg) was randomly fragmented into fragments of approximately 300–350 bp using the NadPrep® DNA universal library construction kit (for MGI) using a Covaris™ fragmentor. After end-repair, A-tailing, and ligation with sequencing adapters, the 300–350 bp fragments were screened using NadPrep® SP Beads. PCR amplification was performed, and the PCR product was purified again using NadPrep® SP Beads to generate a sequencing library. After library construction, initial quantification was performed using Qubit 2.0, followed by detection of inserts using a Bioanalyzer® (Agilent). If the inserts met expectations, pooling was performed based on the library concentration and target data volume. Sequencing was then performed using a DNBSEQ-T7 sequencer using a PE150 sequencing strategy.

[0051] 1.4 Data Collection of Common Horse Breeds Resequencing data from 24 horses, representing two breeds, Thoroughbred and Ili, were downloaded from the NCBI database (Table 1). These two breeds serve as exogenous reference populations. Fastq reads were extracted from the SRA files using SRAtools software.

[0052] Table 1 Resequencing data information of well-known domestic and foreign bred horse breeds

[0053] 1.5 Data filtering and pre-processing A series of quality controls (QC) were performed on the downloaded raw data using FastQ, including removing reads with adapters, removing paired reads with N content exceeding 1% of the total number of bases in the read, and removing paired reads with low-quality (Q ≤ 5) bases exceeding 50% of the total number of bases in the read.

[0054] After data filtering, clean reads were aligned to the reference genome (EquCab3.0) using BWA 0.7.17 (men) software, sorted and indexed using samtools 1.7, and the Bam file was deduplicated using GATK 4.1.8.0 software. The sequencing depth, genome coverage, and other information of each sample were then statistically analyzed based on the Bam file to prepare for subsequent variant detection.

[0055] Based on the alignment of clean reads to the reference genome, SNPs (single nucleotide polymorphisms) were called using GATK 4.1.8.0. SNPs were then strictly filtered using the following criteria: "QD < 2.0 || FS > 60.0 || MQ < 40.0 || SOR > 3.0 || MQRankSum < -12.5 || ReadPosRankSum < -8.0." Functional and positional annotations were performed on the filtered files using SNPeff software.

[0056] 1.6 Inter-population Fst and θ π Ratio population evolution and selection analysis All individuals were subjected to PCA principal component analysis using Plink software, population structure analysis using Admixture software, and phylogenetic tree construction using VCF2DIS software. Based on the above results, all individuals were grouped (see Table 2). After grouping, the Fst value and θ of the improved population and other populations were calculated using VCFtools software. π Calculate θ with a window of 100kb and a step size of 10kb. π Ratio value, select Fst value and θ π The regions with the top 1% Ratio values ​​are selected as candidate regions.

[0057] 1.4 Fst and θ between extreme individuals π Ratio population evolution and selection analysis Sequenced individuals were sorted by body length, and 19 horses in the upper and lower quartiles were selected to construct a maximum body length group (body length >150 cm) and a minimum body length group (body length <140 cm). Fst and θπ values ​​were calculated using vcftools software with a 100 kb window and 10 kb step size. θπ Ratio values ​​were calculated, and regions with the top 1% of Fst and θπ Ratio values ​​were selected as candidate regions associated with body length.

[0058] 1.5 Association analysis with body size traits The intersection of candidate regions between populations and extreme individuals was determined, and SNP markers and their genotypes were extracted from the overlapping candidate regions. The genotype and allele frequencies of each locus were calculated using R software, and differences were analyzed using the chi-square test. Differences in body size traits between individuals with different genotypes were compared using one-way analysis of variance, and the LSD method was used for multiple comparisons of phenotypic values ​​of traits within individuals with different genotypes.

[0059] 2. Test results 2.1 SNP markers associated with body length and chest girth traits in horses A SNP marker rs1136176196 related to the horse body length trait was screened. The site is located at 76,788,230 bp on chromosome 15 of the horse. The two alleles at this site are A / G, and the distribution differences among different populations are significant.

[0060] 2.2 Differentiation and frequency distribution of rs1136176196 locus among populations The inter-group Fst values ​​and θ of 80 horses in 5 groups were used to analyze the π Ratio value results show ( Figure 1 ), the chromosome segment where the rs1136176196 locus is located is selected among different populations ( Fst = 0.10, θ π Ratio = 2.18), indicating that the polymorphism of this site was strongly differentiated among different populations, suggesting that the polymorphism at this site may be an important factor leading to phenotypic differences among different populations.

[0061] The chi-square test results showed that the distribution of each genotype of this SNP marker in different populations was significantly different (Table 2), among which the distribution difference of this site in the first-generation hybrid horse and Kazakh horse and Yili horse was extremely significant ( p <0.01), and the distribution difference between first-generation crossbred horses and purebred horses was significant ( p <0.05), the difference between first-generation hybrid horses and high-generation hybrid horses was not significant ( p >0.05). In first-generation hybrid horses, high-generation hybrid horses, and purebred horses, the AA genotype was the dominant genotype, in Kazakh horses the AG genotype was the dominant genotype, and in Ili horses the frequencies of the three genotypes were similar.

[0062] Table 2 Frequencies of genotypes at rs1136176196 in different populations

[0063] 2.3 Differentiation degree and frequency distribution of rs1136176196 locus among extreme individuals The intergroup Fst values ​​and θ values ​​of 19 horses in two extreme groups were obtained. π Ratio value display ( Figure 2 ), compared with the very short body length group, the chromosome segment where the rs1136176196 locus is located is selected in the very long body length group ( Fst = 0.12, θ π Ratio =7.70), indicating that the polymorphism of this site was strongly differentiated among extreme individuals, suggesting that the polymorphism at this site may be an important factor leading to individual body length differences.

[0064] The chi-square test results showed that the distribution of this SNP marker in the extreme body length group and the extreme body length group was significantly different ( p <0.01). As shown in Table 3, in the extremely large body length group, the AA genotype accounted for 100%, while in the extremely small body length group, the AA genotype accounted for only 57.1%.

[0065] Table 3 Frequencies of different genotypes at rs1136176196 in extreme individuals

[0066] 2.4 Association analysis between rs1136176196 and body size traits The statistical results of seven body size traits (body height, body length, chest circumference, tube circumference, body length ratio, chest circumference ratio and tube circumference ratio) measured in 30 first-generation hybrid horses and 12 high-generation hybrid horses are shown in Table 4.

[0067] Table 4 Statistical results of body size traits

[0068] Association analysis showed the effects of different genotypes of rs1136176196 on seven body size traits, as shown in Table 5. The AA and AG genotypes of rs1136176196 had significant effects on body length and chest circumference of horses in Bortala Prefecture ( p <0.05), with no significant effect on body height, tube circumference, body length ratio, chest circumference ratio, or tube circumference ratio. The average body length and chest circumference of individuals with the AA genotype were greater than those with the AG genotype, with average increases of 5.90% and 4.00%, respectively.

[0069] Table 5 Association analysis between rs1136176196 and body size traits (MEAN+SD)

[0070] Note: Different lowercase letters indicate significant differences between different genotypes of the same trait at the same locus (P<0.05).

[0071] These results indicate that the rs1136176196 mutation can affect body length and chest girth in horses. The AA genotype frequency was 100% in the taller population, significantly higher than in the shorter population. Furthermore, individuals with the AA genotype in the Bortala region had significantly greater body length and chest girth than those with the AG genotype. Typing this SNP marker could predict body length and chest girth in horses, allowing for early selection of horses with slender bodies and larger chest girths. SNP marker-assisted selection could also be used to achieve targeted breeding of trotting racehorses for the 1000 / 5000 m distance, potentially improving the body length and chest girth traits in horses in the Bortala region.

[0072] In the description of this specification, the description with reference to the terms "one embodiment / method", "some embodiments / methods", "example", "specific example" or "some examples" means that the specific features, structures, materials or characteristics described in conjunction with the embodiment / method or example are included in at least one embodiment / method or example of the present invention. In this specification, the schematic expressions of the above terms do not necessarily refer to the same embodiment / method or example. Moreover, the specific features, structures, materials or characteristics described may be combined in an appropriate manner in any one or more embodiments / methods or examples. In addition, those skilled in the art may combine and combine different embodiments / methods or examples described in this specification and the features of different embodiments / methods or examples, unless they are contradictory.

[0073] It should be noted that, in the present invention, relational terms such as "first" and "second" are merely used to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply the existence of any such actual relationship or order between these entities or operations. Moreover, the terms "comprises", "includes" or any other variants thereof are intended to cover non-exclusive inclusion, so that a process, method, article or device comprising a series of elements includes not only those elements, but also other elements not explicitly listed, or also includes elements inherent to such process, method, article or device. In the absence of further restrictions, an element defined by the sentence "comprises a..." does not exclude the presence of other identical elements in the process, method, article or device comprising the element. In the present invention, the meaning of "plurality" is at least two, such as two, three, etc., unless otherwise clearly specified.

[0074] The foregoing description is intended only to provide specific embodiments of the present invention, which will enable those skilled in the art to understand and implement the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention is not intended to be limited to the embodiments shown herein, but is intended to be accorded the widest scope consistent with the principles and novel features claimed herein.

Claims

1. Application of SNP markers associated with horse body length and chest girth traits in any of (1) to (4): (1) Identify or assist in identifying the body length or chest girth of horses; (2) Preparation of products for identifying or assisting in identifying the body length or chest girth traits of horses; (3) Breeding horses with slender bodies and large chest circumference; (4) Preparation of products for breeding horses with slender bodies and large chest circumference; The SNP marker is located at 76,788,230bp of chromosome 15 DNA of the horse genome, the allele is A or G, and the rs ID is rs1136176196.

2. Use of a reagent for detecting SNP markers associated with body length and chest girth traits in any of (1) to (4): (1) Identify or assist in identifying the body length or chest girth of horses; (2) Preparation of products for identifying or assisting in identifying the body length or chest girth traits of horses; (3) Breeding horses with slender bodies and large chest circumference; (4) Preparation of products for breeding horses with slender bodies and large chest circumference; The SNP marker is located at 76,788,230bp of chromosome 15 DNA of the horse genome, the allele is A or G, and the rs ID is rs1136176196.

3. Use of a primer set for amplifying SNP markers associated with body length and chest girth traits in any of (1) to (4): (1) Identify or assist in identifying the body length or chest girth of horses; (2) Preparation of products for identifying or assisting in identifying the body length or chest girth traits of horses; (3) Breeding horses with slender bodies and large chest circumference; (4) Preparation of products for breeding horses with slender bodies and large chest circumference; The SNP marker is located at 76,788,230bp of chromosome 15 DNA of the horse genome, the allele is A or G, and the rs ID is rs1136176196.

4. The use according to any one of claims 1 to 3, characterized in that: The product includes one of a kit, a chip, and an instrument.

5. A method for identifying or assisting in identifying the body length or chest girth traits of a horse, characterized in that: include: Detect the genotypes of SNP markers related to horse body length and chest girth traits in the genome of the test individuals; Predict the body length of the test individual when it grows up based on the genotype: If the genotype of the SNP marker associated with horse body length in the genome of the test individual is the AA genotype, it is predicted that the test individual will grow up to be a horse with a slender body and a large chest circumference; If the genotype of the SNP marker related to horse body length in the genome of the test individual is the AG genotype, it is predicted that the test individual will grow up to be a horse with a short body and a small chest circumference; The SNP marker is located at 76,788,230bp of chromosome 15 DNA of the horse genome, the allele is A or G, and the rs ID is rs1136176196.

6. A method for breeding racehorses, characterized in that: include: Detect the genotypes of SNP markers related to horse body length and chest girth traits in the genome of the test individuals; If the genotype of the SNP marker related to horse body length in the genome of the test individual is AA genotype, the test individual will be bred and managed as a reserve racehorse; If the SNP marker related to horse body length in the genome of the test individual is AG or GG genotype, the test individual will be eliminated; The SNP marker is located at 76,788,230bp of chromosome 15 DNA of the horse genome, the allele is A or G, and the rs ID is rs1136176196.

7. A method for breeding racing stallions, characterized in that: include: Detect the genotypes of SNP markers related to horse body length and chest girth traits in the genome of the test individuals; If the genotype of the SNP marker related to horse body length in the genome of the test individual is AA genotype, the test individual will be bred and managed as a reserve racehorse; If the SNP marker related to horse body length in the genome of the test individual is AG or GG genotype, the test individual will be eliminated; The SNP marker is located at 76,788,230bp of chromosome 15 DNA of the horse genome, the allele is A or G, and the rs ID is rs1136176196.

8. The method according to any one of claims 5 to 7, characterized in that: The method for detecting the genotype of the linked SNP marker associated with the horse chest girth trait in the genome of the test individual is any one of the whole genome sequencing method, PCR method, gene chip method, and SNaPshot method.

9. The method according to any one of claims 5 to 7, characterized in that: The test subject is a foal.

10. The method according to claim 9, characterized in that: The test individuals or their parents are horses from Bozhou area.

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